kth.sePublications
Change search
Refine search result
123 1 - 50 of 118
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Rows per page
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sort
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
Select
The maximal number of hits you can export is 250. When you want to export more records please use the Create feeds function.
  • 1.
    Arruda, Lucas C. M.
    et al.
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden..
    Stikvoort, Arwen
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden..
    Lambert, Melanie
    Karolinska Inst, Dept Med, Ctr Hematol & Regenerat Med, Stockholm, Sweden..
    Jin, Liqing
    Univ Hlth Network, Princess Margaret Canc Ctr, Toronto, ON, Canada..
    Sanchez-Rivera, Laura
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Affinity Proteomics.
    Alves, Renato M. P.
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    de Moura, Tales Rocha
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Biomedical Engineering and Health Systems.
    Mim, Carsten
    KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Biomedical Engineering and Health Systems, Structural Biotechnology.
    Lehmann, Soren
    Karolinska Inst, Dept Med, Ctr Hematol & Regenerat Med, Stockholm, Sweden..
    Axelsson-Robertson, Rebecca
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden.;Karolinska Univ Hosp, Dept Immunol & Transfus Med, Stockholm, Sweden..
    Dick, John E.
    Univ Hlth Network, Princess Margaret Canc Ctr, Toronto, ON, Canada.;Univ Toronto, Dept Mol Genet, Toronto, ON, Canada..
    Mattsson, Jonas
    Univ Hlth Network, Princess Margaret Canc Ctr, Toronto, ON, Canada.;Univ Hlth Network, Princess Margaret Canc Ctr, Gloria & Seymour Epstein Chair Cell Therapy & Tra, Toronto, ON, Canada..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Sci Life Lab, Dept Microbiol Tumor & Cell Biol, Stockholm, Sweden..
    Carlsten, Mattias
    Karolinska Inst, Dept Med, Ctr Hematol & Regenerat Med, Stockholm, Sweden.;Karolinska Univ Hosp, Ctr Cell Therapy & Allogene Stem Cell Transplanta, Stockholm, Sweden..
    Uhlin, Michael
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden.; Karolinska Univ Hosp, Dept Immunol & Transfus Med, Stockholm, Sweden..
    A novel CD34-specific T-cell engager efficiently depletes acute myeloid leukemia and leukemic stem cells in vitro and in vivo2022In: Haematologica, ISSN 0390-6078, E-ISSN 1592-8721, Vol. 107, no 8, p. 1786-1795Article in journal (Refereed)
    Abstract [en]

    Less than a third of patients with acute myeloid leukemia (AML) are cured by chemotherapy and/or hematopoietic stem cell transplantation, highlighting the need to develop more efficient drugs. The low efficacy of standard treatments is associated with inadequate depletion of CD34(+) blasts and leukemic stem cells, the latter a drug-resistant subpopulation of leukemia cells characterized by the CD34(+)CD38(-) phenotype. To target these drug-resistant primitive leukemic cells better, we have designed a CD34/CD3 bi-specific T-cell engager (BTE) and characterized its anti-leukemia potential in vitro, ex vivo and in vivo. Our results show that this CD34-specific BTE induces CD34-dependent T-cell activation and subsequent leukemia cell killing in a dose-dependent manner, further corroborated by enhanced T-cell-mediated killing at the singlecell level. Additionally, the BTE triggered efficient T-cell-mediated depletion of CD34(+) hematopoietic stem cells from peripheral blood stem cell grafts and CD34(+) blasts from AML patients. Using a humanized AML xenograft model, we confirmed that the CD34-specific BTE had in vivo efficacy by depleting CD34(+) blasts and leukemic stem cells without side effects. Taken together, these data demonstrate that the CD34-specific BTE has robust antitumor effects, supporting development of a novel treatment modality with the aim of improving outcomes of patients with AML and myelodysplastic syndromes.

  • 2. Benninger, R. K. P.
    et al.
    Önfelt, Björn
    Neil, M. A. A.
    Davis, D. M.
    French, P. M. W.
    Fluorescence imaging of two-photon linear dichroism: Cholesterol depletion disrupts molecular orientation in cell membranes2005In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 88, no 1, p. 609-622Article in journal (Refereed)
    Abstract [en]

    The plasma membrane of cells is an ordered environment, giving rise to anisotropic orientation and restricted motion of molecules and proteins residing in the membrane. At the same time as being an organized matrix of defined structure, the cell membrane is heterogeneous and dynamic. Here we present a method where we use fluorescence imaging of linear dichroism to measure the orientation of molecules relative to the cell membrane. By detecting linear dichroism as well as fluorescence anisotropy, the orientation parameters are separated from dynamic properties such as rotational diffusion and homo energy transfer ( energy migration). The sensitivity of the technique is enhanced by using two-photon excitation for higher photo-selection compared to single photon excitation. We show here that we can accurately image lipid organization in whole cell membranes and in delicate structures such as membrane nanotubes connecting two cells. The speed of our wide-field imaging system makes it possible to image changes in orientation and anisotropy occurring on a subsecond timescale. This is demonstrated by time-lapse studies showing that cholesterol depletion rapidly disrupts the orientation of a fluorophore located within the hydrophobic region of the cell membrane but not of a surface bound probe. This is consistent with cholesterol having an important role in stabilizing and ordering the lipid tails within the plasma membrane.

  • 3. Benninger, Richard K. P.
    et al.
    Hofmann, Oliver
    Önfelt, Björn
    Munro, Ian
    Dunsby, Chris
    Davis, Daniel M.
    Neil, Mark A. A.
    French, Paul M. W.
    de Mello, Andrew J.
    Fluorescence-lifetime imaging of DNA-dye interactions within continuous-flow microfluidic systems2007In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 46, no 13, p. 2228-2231Article in journal (Refereed)
  • 4. Benninger, Richard K. P.
    et al.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Young, Stephen
    Taner, Sabrina B.
    Culley, Fiona I.
    Schnyder, Tim
    Neil, Mark A. A.
    Wuestner, Daniel
    French, Paul M. W.
    Davis, Daniel M.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Live Cell Linear Dichroism Imaging Reveals Extensive Membrane Ruffling within the Docking Structure of Natural Killer Cell Immune Synapses2009In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 96, no 2, p. L13-L15Article in journal (Refereed)
    Abstract [en]

    We have applied fluorescence imaging of two-photon linear dichroism to measure the subresolution organization of the cell membrane during formation of the activating (cytolytic) natural killer (NK) cell immune synapse (IS). This approach revealed that the NK cell plasma membrane is convoluted into ruffles at the periphery, but not in the center of a mature cytolytic NK cell IS. Time-lapse imaging showed that the membrane ruffles formed at the initial point of contact between NK cells and target cells and then spread radialy across the intercellular contact as the size of the IS increased, becoming absent from the center of the mature synapse. Understanding the role of such extensive membrane ruff ling in the assembly of cytolytic synapses is an intriguing new goal.

  • 5.
    Brandt, Ludwig
    et al.
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics.
    Pfefferle, Aline
    Goodridge, Jodie
    Malmberg, Karl-Johan
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Cytotoxicity and killing kinetics of KIR educated NK cells2017In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 86, no 4, p. 301-301Article in journal (Other academic)
  • 6. Carannante, Valentina
    et al.
    Olofsson, Karl
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    van Ooijen, Hanna
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Edwards, Steven
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Lundqvist, Andreas
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Novel platform for studying infiltration, migration and cytotoxicity of human Natural Killer cells in solid tumors2017In: Scandinavian Journal of Immunology, ISSN 0300-9475, E-ISSN 1365-3083, Vol. 86, no 4, p. 315-315Article in journal (Other academic)
  • 7.
    Carannante, Valentina
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sandström, Niklas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olofsson, Karl
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    van Ooijen, Hanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Hell, Birte
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Wiklund, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. Center for Infectious Medicine, Dept. of Medicine Huddinge, Karolinska Institutet, Stockholm, Sweden.
    Generation of tumor spheroids in microwells to study NK cell cytotoxicity, infiltration and phenotype2023In: Methods in Cell Biology, Elsevier BV , 2023, Vol. 178, p. 195-208Chapter in book (Other academic)
    Abstract [en]

    The development of new immunotherapeutic drugs and combinatorial strategies requires the implementation of novel methods to test their efficacy in vitro. Here, we present a series of miniaturized in vitro assays to assess immune cell cytotoxic activity, infiltration, and phenotype in renal carcinoma spheroids with the use of a recently developed multichambered microwell chip. We provide protocols for tumor spheroid formation, NK cell culture, fluorescence labelling and imaging of live or fixed cells directly in the chip together with data analysis.

  • 8.
    Carannante, Valentina
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Wiklund, Martin
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Sci Life Lab, Stockholm, Sweden..
    In vitro models to study natural killer cell dynamics in the tumor microenvironment2023In: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 14, article id 1135148Article, review/survey (Refereed)
    Abstract [en]

    Immunotherapy is revolutionizing cancer therapy. The rapid development of new immunotherapeutic strategies to treat solid tumors is posing new challenges for preclinical research, demanding novel in vitro methods to test treatments. Such methods should meet specific requirements, such as enabling the evaluation of immune cell responses like cytotoxicity or cytokine release, and infiltration into the tumor microenvironment using cancer models representative of the original disease. They should allow high-throughput and high-content analysis, to evaluate the efficacy of treatments and understand immune-evasion processes to facilitate development of new therapeutic targets. Ideally, they should be suitable for personalized immunotherapy testing, providing information for patient stratification. Consequently, the application of in vitro 3-dimensional (3D) cell culture models, such as tumor spheroids and organoids, is rapidly expanding in the immunotherapeutic field, coupled with the development of novel imaging-based techniques and -omic analysis. In this paper, we review the recent advances in the development of in vitro 3D platforms applied to natural killer (NK) cell-based cancer immunotherapy studies, highlighting the benefits and limitations of the current methods, and discuss new concepts and future directions of the field.

  • 9.
    Christakou, Athanasia E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Kadri, N.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. Karolinska Institute, Sweden.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Characterization of natural killer cells' cytotoxic heterogeneity using an array of sono-cages2012In: Proceedings of the 16th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2012, Chemical and Biological Microsystems Society , 2012, p. 1555-1557Conference paper (Refereed)
    Abstract [en]

    Using a multi-well device as an array of sono-cages for single cell analysis, we quantify for the first time the heterogeneity of natural killer (NK) cells' cytotoxic response against cancer cells. We report a fraction of inactive NK cells within the tested population (36%), as well as the existence of few 'serial killers' that eliminate up to six targets during 4 hours. We also characterize the multi-well acoustic device in terms of trapping efficiency at different actuation voltages, using adherent and non-adherent cell lines. We show that the acoustic forces applied on the cells can be compared to forces of biological processes (i.e. cell adherence).

  • 10.
    Christakou, Athanasia. E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Khorshidi, Mohammad Ali
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Aggregation and long-term positioning of cells by ultrasound in a multi-well microchip for high-resolution imaging of the natural killer cell immune synapse2011In: 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences 2011, MicroTAS 2011, 2011, p. 329-331Conference paper (Refereed)
    Abstract [en]

    In this study we investigate the ability of Natural Killer (NK) cells to form ultrasound-mediated intercellular contacts with target cells in a multi-well microdevice by high-resolution confocal-microscopy imaging of inhibitory immune synapses. Furthermore, we compare the NK-Target cell cluster migration with and without ultrasound actuation. Experiments indicate that clusters of cells are positioned and maintained centered in the wells for 17 hours when they are exposed continuously to ultrasound. Our system can be used for both screening high numbers of events in low resolution and also for high resolution imaging of long term cell-cell interactions.

  • 11.
    Christakou, Athanasia E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Khorshidi, Mohammad Ali
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Kadri, Nadir
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Live cell imaging in a micro-array of acoustic traps facilitates quantification of natural killer cell heterogeneity2013In: Integrative Biology, ISSN 1757-9694, E-ISSN 1757-9708, Vol. 5, no 4, p. 712-719Article in journal (Refereed)
    Abstract [en]

    Natural killer (NK) cells kill virus-infected or cancer cells through the release of cytotoxic granules into a tight intercellular contact. NK cell populations comprise individual cells with varying sensitivity to distinct input signals, leading to disparate responses. To resolve this NK cell heterogeneity, we have designed a novel assay based on ultrasound-assisted cell-cell aggregation in a multiwell chip allowing high-resolution time-lapse imaging of one hundred NK-target cell interactions in parallel. Studying human NK cells' ability to kill MHC class I deficient tumor cells, we show that approximately two thirds of the NK cells display cytotoxicity, with some NK cells being particularly active, killing up to six target cells during the assay. We also report that simultaneous interaction with several susceptible target cells increases the cytotoxic responsiveness of NK cells, which could be coupled to a previously unknown regulatory mechanism with implications for NK-mediated tumor elimination.

  • 12.
    Christakou, Athanasia E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Solid tumor spheroid formation by temperature-controlled high voltage ultrasound in a multi-well microdevice2014In: 18th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2014, Chemical and Biological Microsystems SocietyChemical and Biological Microsystems Society , 2014, p. 573-575Conference paper (Refereed)
    Abstract [en]

    In the present work we demonstrate effective 3D growth of human hepatocellular carcinoma (HCC) HepG2 cell spheroids in parallel in a multi-well microdevice actuated with high voltage ultrasound in a temperature-controlled system. We compare the spheroid formation during continuous ultrasound exposure for one week where we formed spheroids in 59% of the wells, with the spheroid formation without ultrasound actuation, where we obtained 0% spheroids. Furthermore, we present an application of the tumor spheroids for investigating natural killer (NK) cells behavior against solid tumors.

  • 13.
    Christakou, Athanasia E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ultrasonic three-dimensional cell culture on chip for dynamic studies of tumor immune surveillance by natural killer cellsManuscript (preprint) (Other academic)
  • 14.
    Christakou, Athanasia E.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ultrasound-assisted three-dimensional tumor formation in a multi well microplate for monitoring natural killer cell functional behaviorManuscript (preprint) (Other academic)
  • 15.
    Christakou, Athanasia
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Characterization of natural killer cell immune surveillance against solid liver tumors2015In: MicroTAS 2015 - 19th International Conference on Miniaturized Systems for Chemistry and Life Sciences, Chemical and Biological Microsystems Society , 2015, p. 915-917Conference paper (Refereed)
    Abstract [en]

    We demonstrate a method for investigating natural killer (NK) cell aggression against ultrasound-assisted human hepatocellular carcinoma (HCC) HepG2 solid tumors in a multi-well microdevice. We quantify the activity of human primary IL-2 activated NK cells against HepG2 tumors for up to five days and we present the correlation between NK cell numbers versus average tumor volume and final tumor outcome (growth or defeat). We suggest future applications on formation of tumors originated from primary tumors cells and other tumor components as well as primary NK originating from the patient for use in personalized immunotherapy.

  • 16.
    Christakou, Athanasia
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ohlin, Mathias
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. Karolinska Inst, Dept Microbiol Tumor & Cell Biol, Stockholm, Sweden.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Ultrasonic three-dimensional on-chip cell culture for dynamic studies of tumor immune surveillance by natural killer cells2015In: Lab on a Chip, ISSN 1473-0197, E-ISSN 1473-0189, Vol. 15, no 15, p. 3222-31Article in journal (Refereed)
    Abstract [en]

    We demonstrate a simple method for three-dimensional (3D) cell culture controlled by ultrasonic standing waves in a multi-well microplate. The method gently arranges cells in a suspension into a single aggregate in each well of the microplate and, by this, nucleates 3D tissue-like cell growth for culture times between two and seven days. The microplate device is compatible with both high-resolution optical microscopy and maintenance in a standard cell incubator. The result is a scaffold- and coating-free method for 3D cell culture that can be used for controlling the cellular architecture, as well as the cellular and molecular composition of the microenvironment in and around the formed cell structures. We demonstrate the parallel production of one hundred synthetic 3D solid tumors comprising up to thousands of human hepatocellular carcinoma (HCC) HepG2 cells, we characterize the tumor structure by high-resolution optical microscopy, and we monitor the functional behavior of natural killer (NK) cells migrating, docking and interacting with the tumor model during culture. Our results show that the method can be used for determining the collective ability of a given number of NK cells to defeat a solid tumor having a certain size, shape and composition. The ultrasound-based method itself is generic and can meet any demand from applications where it is advantageous to monitor cell culture from production to analysis of 3D tissue or tumor models using microscopy in one single microplate device.

  • 17. Coates, C. G.
    et al.
    Olofsson, J.
    Coletti, M.
    McGarvey, J. J.
    Önfelt, Björn
    Lincoln, P.
    Norden, B.
    Tuite, E.
    Matousek, P.
    Parker, A. W.
    Picosecond time-resolved resonance Raman probing of the light-switch states of Ru(Phen)(2)dppz (2+)2001In: Journal of Physical Chemistry B, ISSN 1520-6106, E-ISSN 1520-5207, Vol. 105, no 50, p. 12653-12664Article in journal (Refereed)
    Abstract [en]

    Picosecond time-resolved resonance Raman (picosecond-TR3) spectroscopy has been used to conduct an extensive photophysical characterization of the light- switch complex [Ru(phen)(2)dppz](2+) as a function of environment, in which studies have been carried out in aqueous and nonaqueous media and in DNA. The results are considered in rotation to a previous report describing environment-sensitive lowest triplet MLCT states. Vibrational marker features and enhancement patterns were used to determine the rapid progression (< 20 ps) between two triplet MLCT states in aqueous environment, followed by subnanosecond, nonradiative deactivation to the ground state. In nonaqueous environment, the long-lived, emissive triplet MLCT state is spectrally identified as the short-lived first triplet MLCT state observed in water, in agreement with the earlier proposed mechanism. The present data are shown to correlate well with previous nanosecond RR findings for the complex in each environment. Interestingly, a precursor state has been identified upon excitation in both nonaqueous solvent and in DNA, which precedes the triplet MLCT state, and the lifetime of which appears to be environment dependent. Observation of this state is discussed in relation to other recent femtosecond spectroscopic studies on this complex.

  • 18. Dunsby, C.
    et al.
    Lanigan, P. M. P.
    McGinty, J.
    Elson, D. S.
    Requejo-Isidro, J.
    Munro, I.
    Galletly, N.
    McCann, F.
    Treanor, B.
    Önfelt, Björn
    Davis, D. M.
    Neil, M. A. A.
    French, P. M. W.
    An electronically tunable ultrafast laser source applied to fluorescence imaging and fluorescence lifetime imaging microscopy2004In: Journal of Physics D: Applied Physics, ISSN 0022-3727, E-ISSN 1361-6463, Vol. 37, no 23, p. 3296-3303Article in journal (Refereed)
    Abstract [en]

    Fluorescence imaging is used widely in microscopy and macroscopic imaging applications for fields ranging from biomedicine to materials science. A critical component for any fluorescence imaging system is the excitation source. Traditionally, wide-field systems use filtered thermal or arc-generated white light sources, while point scanning confocal microscope systems require spatially coherent (point-like) laser sources. Unfortunately, the limited range of visible wavelengths available from conventional laser sources constrains the design and usefulness of fluorescent probes in confocal microscopy. A 'hands-off' laser-like source, electronically tunable across the visible spectrum, would be invaluable for fluorescence imaging and provide new opportunities, e.g. automated excitation fingerprinting and in situ measurement of excitation cross-sections. Yet more information can be obtained using fluorescence lifetime imaging (FLIM), which requires that the light source be pulsed or rapidly modulated. We show how a white light continuum, generated by injecting femtosecond optical radiation into a micro-structured optical fibre, coupled with a simple prism-based tunable filter arrangement, can fulfil all these roles as a continuously electronically tunable (435-1150 nm) visible ultrafast light source in confocal, wide-field and FLIM systems.

  • 19.
    Dunst, Josefine
    et al.
    Department of Medicine, Division of Immunology and Allergy, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.
    Glaros, Vassilis
    Department of Medicine, Division of Immunology and Allergy, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.
    Englmaier, Lukas
    Department of Medicine, Division of Immunology and Allergy, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.
    Sandoz, Patrick
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics. Department of Microbiology, Tumor and Cell Biology, Karolinska Institute, Solna, Sweden.
    Kisielow, Jan
    Institute of Molecular Health Sciences, ETH, Zurich, Switzerland.
    Kreslavsky, Taras
    Department of Medicine, Division of Immunology and Allergy, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.
    Recognition of synthetic polyanionic ligands underlies “spontaneous” reactivity of Vγ1 γδTCRs2020In: Journal of Leukocyte Biology, ISSN 0741-5400, E-ISSN 1938-3673, Vol. 107, no 6, p. 1033-1044Article in journal (Refereed)
    Abstract [en]

    Although γδTCRs were discovered more than 30 yr ago, principles of antigen recognition by these receptors remain unclear and the nature of these antigens is largely elusive. Numerous studies reported that T cell hybridomas expressing several Vγ1‐containing TCRs, including the Vγ1Vδ6 TCR of γδNKT cells, spontaneously secrete cytokines. This property was interpreted as recognition of a self‐ligand expressed on the hybridoma cells themselves. Here, we revisited this finding using a recently developed reporter system and live single cell imaging. We confirmed strong spontaneous signaling by Vγ1Vδ6 and related TCRs, but not by TCRs from several other γδ or innate‐like αβ T cells, and demonstrated that both γ and δ chains contributed to this reactivity. Unexpectedly, live single cell imaging showed that activation of this signaling did not require any interaction between cells. Further investigation revealed that the signaling is instead activated by interaction with negatively charged surfaces abundantly present under regular cell culture conditions and was abrogated when noncharged cell culture vessels were used. This mode of TCR signaling activation was not restricted to the reporter cell lines, as interaction with negatively charged surfaces also triggered TCR signaling in ex vivo Vγ1 γδ T cells. Taken together, these results explain long‐standing observations on the spontaneous reactivity of Vγ1Vδ6 TCR and demonstrate an unexpected antigen presentation‐independent mode of TCR activation by a spectrum of chemically unrelated polyanionic ligands.

  • 20.
    Edwards, Steven J.
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Carannante, Valentina
    Karolinska Inst, Sci Life Lab, Dept Microbiol Tumor & Cell Biol, Stockholm, Sweden..
    Kuhnigk, Kyra
    Karolinska Inst, Dept Med Huddinge, Ctr Hematol & Regenerat Med, Stockholm, Sweden..
    Ring, Henrik
    Uppsala Univ, BMC, Dept Neurosci, Uppsala, Sweden..
    Tararuk, Tatsiana
    Uppsala Univ, BMC, Dept Neurosci, Uppsala, Sweden..
    Hallböök, Finn
    Uppsala Univ, BMC, Dept Neurosci, Uppsala, Sweden..
    Blom, Hans
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    High-Resolution Imaging of Tumor Spheroids and Organoids Enabled by Expansion Microscopy2020In: Frontiers in Molecular Biosciences, E-ISSN 2296-889X, Vol. 7, article id 208Article in journal (Refereed)
    Abstract [en]

    Three-dimensional cell cultures are able to better mimic the physiology and cellular environments found in tissuesin vivocompared to cells grown in two dimensions. In order to study the structure and function of cells in 3-D cultures, light microscopy is frequently used. The preparation of 3-D cell cultures for light microscopy is often destructive, including physical sectioning of the samples, which can result in the loss of 3-D information. In order to probe the structure of 3-D cell cultures at high resolution, we have explored the use of expansion microscopy and compared it to a simple immersion clearing protocol. We provide a practical method for the study of spheroids, organoids and tumor-infiltrating immune cells at high resolution without the loss of spatial organization. Expanded samples are highly transparent, enabling high-resolution imaging over extended volumes by significantly reducing light scatter and absorption. In addition, the hydrogel-like nature of expanded samples enables homogenous antibody labeling of dense epitopes throughout the sample volume. The improved labeling and image quality achieved in expanded samples revealed details in the center of the organoid which were previously only observable following serial sectioning. In comparison to chemically cleared spheroids, the improved signal-to-background ratio of expanded samples greatly improved subsequent methods for image segmentation and analysis.

  • 21. Eleme, K.
    et al.
    Taner, S. B.
    Önfelt, Björn
    Collinson, L. M.
    McCann, F. E.
    Chalupny, N. J.
    Cosman, D.
    Hopkins, C.
    Magee, A. I.
    Davis, D. M.
    Cell surface organization of stress-inducible proteins ULBP and MICA that stimulate human NK cells and T cells via NKG2D2004In: Journal of Experimental Medicine, ISSN 0022-1007, E-ISSN 1540-9538, Vol. 199, no 7, p. 1005-1010Article in journal (Refereed)
    Abstract [en]

    Cell surface proteins major histocompatibility complex (MHC) class I-related chain A (MICA) and UL16-binding proteins (ULBP) 1, 2, and 3 are up-regulated upon infection or tumor transformation and can activate human natural killer (NK) cells. Patches of cross-linked raft resident ganglioside GM1 colocalized with ULBP1, 2, 3, or MICA, but not CD45. Thus, ULBPs and MICA are expressed in lipid rafts at the cell surface. Western blotting revealed that glycosylphosphatidylinositol (GPI)-anchored ULBP3 but not transmembrane MICA, MHC class I protein, or transferrin receptor, accumulated in detergent-resistant membranes containing GM1. Thus, MICA may have a weaker association with lipid rafts than ULBP3, yet both proteins accumulate at an activating human N-K cell immune synapse. Target cell lipid rafts marked by green fluorescent protein-tagged GPI also accumulate with ULBP3 at some synapses. Electron microscopy reveals constitutive clusters of ULBP at the cell surface. Regarding a specific molecular basis for the organization of these proteins, ULBP1, 2, and 3 and MICA are lipid modified. ULBP1, 2, and 3 are GPI anchored, and we demonstrate here that MICA is S-acylated. Finally, expression of a truncated form of MICA that lacks the putative site for S-acylation and the cytoplasmic tall can be expressed at the cell surface, but is unable to activate NK cells.

  • 22. Enqvist, M.
    et al.
    Ask, E. H.
    Forslund, E.
    Carlsten, M.
    Abrahamsen, G.
    Béziat, V.
    Andersson, S.
    Schaffer, M.
    Spurkland, A.
    Bryceson, Y.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Malmberg, K. -J
    Coordinated expression of DNAM-1 and LFA-1 in educated NK cells2015In: Journal of Immunology, ISSN 0022-1767, E-ISSN 1550-6606, Vol. 194, no 9, p. 4518-4527Article in journal (Refereed)
    Abstract [en]

    The functional capacity of NK cells is dynamically tuned by integrated signals from inhibitory and activating cell surface receptors in a process termed NK cell education. However, the understanding of the cellular and molecular mechanisms behind this functional tuning is limited. In this study, we show that the expression of the adhesion molecule and activation receptor DNAX accessory molecule 1 (DNAM-1) correlates with the quantity and quality of the inhibitory input by HLA class I-specific killer cell Ig-like receptors and CD94/NKG2A as well as with the magnitude of functional responses. Upon target cell recognition, the conformational state of LFA-1 changed in educated NK cells, associated with rapid colocalization of both active LFA-1 and DNAM-1 at the immune synapse. Thus, the coordinated expression of LFA-1 and DNAM-1 is a central component of NK cell education and provides a potential mechanism for controlling cytotoxicity by functionally mature NK cells.

  • 23.
    Felices, Martin
    et al.
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA..
    Sarhan, Dhifaf
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA.;Karolinska Inst, Dept Oncol Pathol, Stockholm, Sweden..
    Brandt, Ludwig
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH Royal Inst Technol, Dept Appl Phys, Sci Life Lab, Stockholm, Sweden..
    Guldevall, Karolin
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    McElmurry, Ron
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA..
    Lenvik, Alexander
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA..
    Chu, Sami
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA..
    Tolar, Jakub
    Univ Minnesota, Dept Pediat, Div Blood & Marrow Transplantat, Minneapolis, MN 55455 USA.;Univ Minnesota, Stem Cell Inst, Minneapolis, MN USA..
    Taras, Elizabeth
    Univ Minnesota, Dept Therapeut Radiol Radiat Oncol, Lab Mol Canc Therapeut, Masonic Canc Ctr, Minneapolis, MN USA..
    Spellman, Stephen R.
    Natl Marrow Donor Program, Minneapolis, MN USA..
    Warlick, Erica D.
    Univ Minnesota, Hematol Oncol & Transplantat, Minneapolis, MN USA..
    Verneris, Michael R.
    Univ Minnesota, Dept Pediat, Div Pediat Hematol Oncol Bone & Marrow Transplant, Minneapolis, MN 55455 USA..
    Cooley, Sarah
    Univ Minnesota, Dept Med, Div Hematol Oncol & Transplantat, Box 736 UMHC, Minneapolis, MN 55455 USA..
    Weisdorf, Daniel
    Univ Minnesota, Hematol Oncol & Transplantat, Minneapolis, MN USA..
    Blazar, Bruce R.
    Univ Minnesota, Dept Pediat, Blood & Marrow Transplantat, Minneapolis, MN 55455 USA..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Vallera, Dan
    Univ Minnesota, Dept Therapeut Radiol Radiat Oncol, Masonic Canc Ctr, Minneapolis, MN USA..
    Miller, Jeffrey S.
    Univ Minnesota, Hematol Oncol & Transplantat, Minneapolis, MN USA..
    CD16-IL15-CD33 Trispecific Killer Engager (TriKE) Overcomes Cancer-Induced Immune Suppression and Induces Natural Killer Cell-Mediated Control of MDS and AML Via Enhanced Killing Kinetics2016In: Blood, ISSN 0006-4971, E-ISSN 1528-0020, Vol. 128, no 22Article in journal (Other academic)
  • 24.
    Fleming, Cassandra L.
    et al.
    Chalmers Univ Technol, Dept Chem & Chem Engn Phys Chem, S-41296 Gothenburg, Sweden.;Univ Gothenburg, Dept Chem & Mol Biol, S-41296 Gothenburg, Sweden..
    Sandoz, Patrick A.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Inghardt, Tord
    AstraZeneca, Med Chem, Res & Early Dev Cardiovasc Renal & Metab, BioPharmaceut R&D, Gothenburg, Sweden..
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Grotli, Morten
    Univ Gothenburg, Dept Chem & Mol Biol, S-41296 Gothenburg, Sweden..
    Andreasson, Joakim
    Chalmers Univ Technol, Dept Chem & Chem Engn Phys Chem, S-41296 Gothenburg, Sweden..
    A Fluorescent Kinase Inhibitor that Exhibits Diagnostic Changes in Emission upon Binding2019In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773Article in journal (Refereed)
    Abstract [en]

    The development of a fluorescent LCK inhibitor that exhibits favourable solvatochromic properties upon binding the kinase is described. Fluorescent properties were realised through the inclusion of a prodan-derived fluorophore into the pharmacophore of an ATP-competitive kinase inhibitor. Fluorescence titration experiments demonstrate the solvatochromic properties of the inhibitor, in which dramatic increase in emission intensity and hypsochromic shift in emission maxima are clearly observed upon binding LCK. Microscopy experiments in cellular contexts together with flow cytometry show that the fluorescence intensity of the inhibitor correlates with the LCK concentration. Furthermore, multiphoton microscopy experiments demonstrate both the rapid cellular uptake of the inhibitor and that the two-photon cross section of the inhibitor is amenable for excitation at 700 nm.

  • 25.
    Fogelqvist, Emelie
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Kördel, Mikael
    Carannante, Valentina
    Önfelt, Björn
    Hertz, Hans
    Laboratory cryo x-ray microscopy for 3D cell imagingManuscript (preprint) (Other academic)
  • 26.
    Fogelqvist, Emelie
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Kördel, Mikael
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Carannante, Valentina
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cellular Biophysics.
    Hertz, Hans
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics. Karolinska Institutet, Sweden.
    Laboratory cryo x-ray microscopy for 3D cell imaging2017In: Scientific Reports, E-ISSN 2045-2322, Vol. 7, article id 13433Article in journal (Refereed)
    Abstract [en]

    Water-window x-ray microscopy allows two-and three-dimensional (2D and 3D) imaging of intact unstained cells in their cryofixed near-native state with unique contrast and high resolution. Present operational biological water-window microscopes are based at synchrotron facilities, which limits their accessibility and integration with complementary methods. Laboratory-source microscopes have had difficulty addressing relevant biological tasks with proper resolution and contrast due to long exposure times and limited up-time. Here we report on laboratory cryo x-ray microscopy with the exposure time, contrast, and reliability to allow for routine high-spatial resolution 3D imaging of intact cells and cell-cell interactions. Stabilization of the laser-plasma source combined with new optics and sample preparation provide high-resolution cell imaging, both in 2D with ten-second exposures and in 3D with twenty-minute tomography. Examples include monitoring of the distribution of carbon-dense vesicles in starving HEK293T cells and imaging the interaction between natural killer cells and target cells.

  • 27. Forslund, E.
    et al.
    Guldevall, Karolin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Olofsson, Per E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Christakou, Athanasia E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Novel microchip-based tools facilitating live cell imaging and assessment of functional heterogeneity within NK cell populations2012In: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 3, no OCT, p. 300-Article in journal (Refereed)
    Abstract [en]

    Each individual has a heterogeneous pool of NK cells consisting of cells that may be specialized towards specific functional responses such as secretion of cytokines or killing of tumor cells. Many conventional methods are not fit to characterize heterogeneous populations as they measure the average response of all cells. Thus, there is a need for experimental platforms that provide single cell resolution. In addition, there are transient and stochastic variations in functional responses at the single cell level, calling for methods that allow studies of many events over extended periods of time. This paper presents a versatile microchip platform enabling long-term microscopic studies of individual NK cells interacting with target cells. Each microchip contains an array of microwells, optimized for medium or high-resolution time-lapse imaging of single or multiple NK and target cells, or for screening of thousands of isolated NK-target cell interactions. Individual NK cells confined with target cells in small microwells is a suitable setup for high-content screening and rapid assessment of heterogeneity within populations, while microwells of larger dimensions are appropriate for studies of NK cell migration and sequential interactions with multiple target cells. By combining the chip technology with ultrasonic manipulation, NK and target cells can be forced to interact and positioned with high spatial accuracy within individual microwells.This setup effectively and synchronously creates NK-target conjugates at hundreds of parallel positions in the microchip. Thus, this facilitates assessment of temporal aspects of NK-target cell interactions, e.g., conjugation, immune synapse formation, and cytotoxic events.The microchip platform presented here can be used to effectively address questions related to fundamental functions of NK cells that can lead to better understanding of how the behavior of individual cells add up to give a functional response at the population level.

  • 28. Forslund, Elin
    et al.
    Sohlberg, Ebba
    Enqvist, Monika
    Olofsson, Per E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Malmberg, Karl-Johan
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Sci Life Lab, Dept Microbiol Tumor & Cell Biol, S-17165 Stockholm, Sweden.
    Microchip-Based Single-Cell Imaging Reveals That CD56(dim) CD57(-)KIR(-)NKG2A(+) NK Cells Have More Dynamic Migration Associated with Increased Target Cell Conjugation and Probability of Killing Compared to CD56(dim)CD57(-)KIR(-)NKG2A(-) NK Cells2015In: Journal of Immunology, ISSN 0022-1767, E-ISSN 1550-6606, Vol. 195, no 7, p. 3374-3381Article in journal (Refereed)
    Abstract [en]

    NK cells are functionally educated by self-MHC specific receptors, including the inhibitory killer cell Ig-like receptors (KIRs) and the lectin-like CD94/NKG2A heterodimer. Little is known about how NK cell education influences qualitative aspects of cytotoxicity such as migration behavior and efficacy of activation and killing at the single-cell level. In this study, we have compared the behavior of FACS-sorted CD56(dim)CD57(-)KIR(-)NKG2A(+) (NKG2A(+)) and CD56(dim)CD57(-)KIR(-)NKG2A(+) (lacking inhibitory receptors; IR-) human NK cells by quantifying migration, cytotoxicity, and contact dynamics using microchip-based live cell imaging. NKG2A(+) NK cells displayed a more dynamic migration behavior and made more contacts with target cells than IR-NK cells. NKG2A(+) NK cells also more frequently killed the target cells once a conjugate had been formed. NK cells with serial killing capacity were primarily found among NKG2A(+) NK cells. Conjugates involving IR- NK cells were generally more short-lived and IR- NK cells did not become activated to the same extent as NKG2A(+) NK cells when in contact with target cells, as evident by their reduced spreading response. In contrast, NKG2A(+) and IR- NK cells showed similar dynamics in terms of duration of conjugation periods and NK cell spreading response in conjugates that led to killing. Taken together, these observations suggest that the high killing capacity of NKG2A(+) NK cells is linked to processes regulating events in the recognition phase of NK-target cell contact rather than events after cytotoxicity has been triggered.

  • 29. Frisk, T.
    et al.
    Guldevall, Karolin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Vanherbergen, B.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Live-cell imaging of natural killer cell mediated tumor rejection in arrays of microwells2010In: 14th International Conference on Miniaturized Systems for Chemistry and Life Sciences 2010, MicroTAS 2010: Volume 2, 2010, p. 950-952Conference paper (Refereed)
    Abstract [en]

    Due to the inherent heterogeneity of immune cell populations a comprehensive view of immune functions can only be achieved by collecting data from many individual cells. We here demonstrate a novel multi-well microchip platform for cell analysis, which we use to study the interaction between natural killer (NK) cells and their target cells.

  • 30.
    Frisk, Thomas
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Khorshidi, Mohammad Ali
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Guldevall, Karolin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    A silicon-glass microwell platform for high-resolution imaging and high-content screening with single cell resolution2011In: Biomedical microdevices (Print), ISSN 1387-2176, E-ISSN 1572-8781, Vol. 13, no 4, p. 683-693Article in journal (Refereed)
    Abstract [en]

    We present a novel microwell array platform suited for various cell-imaging assays where single cell resolution is important. The platform consists of an exchangeable silicon-glass microchip for cell biological applications and a custom made holder that fits in conventional microscopes. The microchips presented here contain arrays of miniature wells, where the well sizes and layout have been designed for different applications, including single cell imaging, studies of cell-cell interactions or ultrasonic manipulation of cells. The device has been designed to be easy to use, to allow long-term assays (spanning several days) with read-outs based on high-resolution imaging or high-content screening. This study is focused on screening applications and an automatic cell counting protocol is described and evaluated. Finally, we have tested the device and automatic counting by studying the selective survival and clonal expansion of 721.221 B cells transfected to express HLA Cw6-GFP compared to untransfected 721.221 B cells when grown under antibiotic selection for 3 days. The device and automated analysis protocol make up the foundation for development of several novel cellular imaging assays.

  • 31.
    Gaballa, Ahmed
    et al.
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden..
    Stikvoort, Arwen
    Karolinska Inst, Dept Oncol & Pathol, Stockholm, Sweden..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Microbiol Tumor & Cell Biol, Stockholm, Sweden..
    Mattsson, Jonas
    Karolinska Inst, Dept Oncol & Pathol, Stockholm, Sweden..
    Sundin, Mikael
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden.;Karolinska Univ Hosp, Astrid Lindgren Childrens Hosp, Sect Paediat Hematol Immunol & Hematopoiet Cell T, Stockholm, Sweden..
    Watz, Emma
    Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden.;Karolinska Univ Hosp, Dept Immunol & Transfus Med, Stockholm, Sweden..
    Uhlin, Michael
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Clin Sci Intervent & Technol, Stockholm, Sweden.;Karolinska Univ Hosp, Dept Immunol & Transfus Med, Stockholm, Sweden..
    T-cell frequencies of CD8(+) gamma delta and CD27(+) gamma delta cells in the stem cell graft predict the outcome after allogeneic hematopoietic cell transplantation2019In: Bone Marrow Transplantation, ISSN 0268-3369, E-ISSN 1476-5365, Vol. 54, no 10, p. 1562-1574Article in journal (Refereed)
    Abstract [en]

    The impact of intra-graft T cells on the clinical outcome after allogeneic hematopoietic cell transplantation has been investigated. Most previous studies have focused on the role of alpha beta cells while gamma delta cells have received less attention. It has been an open question whether gamma delta cells are beneficial or not for patient outcome, especially with regards to graft versus host disease. In this study, graft composition of.d cell subsets was analyzed and correlated to clinical outcome in 105 recipients who underwent allogeneic hematopoietic cell transplantation between 2013 and 2016. We demonstrate for the first time that grafts containing higher T-cell proportions of CD8(+) gamma delta cells were associated with increased cumulative incidence of acute graft versus host disease grade II-III (50% vs 22.6%; P = 0.008). Additionally, graft T-cell frequency of CD27(+) gamma delta cells was inversely correlated with relapse (P = 0.006) and CMV reactivation (P = 0.05). We conclude that clinical outcome after allogeneic hematopoietic cell transplantation is influenced by the proportions of distinct gamma delta cell subsets in the stem cell graft. We also provide evidence that CD8(+) gamma delta cells are potentially alloreactive and may play a role in acute graft versus host disease. This study illustrates the importance of better understanding of the role of distinct subsets of.d cells in allogeneic hematopoietic cell transplantation.

  • 32.
    Girnyk, Maksym A.
    et al.
    KTH, School of Electrical Engineering (EES), Communication Theory.
    Vehkapera, Mikko
    Rasmussen, Lars Kildehoj
    Christakou, Athanasia
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Onfelt, Bjorn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Orange, Jordan
    Lytic granule convergence is essential for NK cells to promote targeted killing while preventing collateral damage2016In: Journal of Immunology, ISSN 0022-1767, E-ISSN 1550-6606, Vol. 196Article in journal (Other academic)
  • 33. Goodridge, Jodie P.
    et al.
    Onfelt, Bjorn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. Karolinska Inst, Microbiol Tumor & Cell Biol.
    Malmberg, Karl-Johan
    Newtonian cell interactions shape natural killer cell education2015In: Immunological Reviews, ISSN 0105-2896, E-ISSN 1600-065X, Vol. 267, no 1, p. 197-213Article, review/survey (Refereed)
    Abstract [en]

    Newton's third law of motion states that for every action on a physical object there is an equal and opposite reaction. The dynamic change in functional potential of natural killer (NK) cells during education bears many features of such classical mechanics. Cumulative physical interactions between cells, under a constant influence of homeostatic drivers of differentiation, lead to a reactive spectrum that ultimately shapes the functionality of each NK cell. Inhibitory signaling from an array of self-specific receptors appear not only to suppress self-reactivity but also aid in the persistence of effector functions over time, thereby allowing the cell to gradually build up a functional potential. Conversely, the frequent non-cytolytic interactions between normal cells in the absence of such inhibitory signaling result in continuous stimulation of the cells and attenuation of effector function. Although an innate cell, the degree to which the fate of the NK cell is predetermined versus its ability to adapt to its own environment can be revealed through a Newtonian view of NK cell education, one which is both chronological and dynamic. As such, the development of NK cell functional diversity is the product of qualitatively different physical interactions with host cells, rather than simply the sum of their signals or an imprint based on intrinsically different transcriptional programs.

  • 34.
    Guldevall, Karolin
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Brandt, Ludwig
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Forslund, Elin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Microbiol Tumor & Cell Biol, Sweden.
    Olofsson, Karl
    Frisk, Thomas W.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olofsson, Per E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gustafsson, Karin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Manneberg, Otto
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Karre, Klas
    Uhlin, Michael
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Microchip screening Platform for single cell assessment of NK cell cytotoxicity2016In: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 7, article id 119Article in journal (Refereed)
    Abstract [en]

    Here, we report a screening platform for assessment of the cytotoxic potential of individual natural killer (NK) cells within larger populations. Human primary NK cells were distributed across a silicon-glass microchip containing 32,400 individual microwells loaded with target cells. Through fluorescence screening and automated image analysis, the numbers of NK and live or dead target cells in each well could be assessed at different time points after initial mixing. Cytotoxicity was also studied by time-lapse live-cell imaging in microwells quantifying the killing potential of individual NK cells. Although most resting NK cells (approximate to 75%) were non-cytotoxic against the leukemia cell line K562, some NK cells were able to kill several (>= 3) target cells within the 12-h long experiment. In addition, the screening approach was adapted to increase the chance to find and evaluate serial killing NK cells. Even if the cytotoxic potential varied between donors, it was evident that a small fraction of highly cytotoxic NK cells were responsible for a substantial portion of the killing. We demonstrate multiple assays where our platform can be used to enumerate and characterize cytotoxic cells, such as NK or T cells. This approach could find use in clinical applications, e.g., in the selection of donors for stem cell transplantation or generation of highly specific and cytotoxic cells for adoptive immunotherapy.

  • 35.
    Guldevall, Karolin
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Khorsidi, Mohammed Ali
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Manneberg, Otto
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Christakou, Athanasia
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Imaging immune surveillance by individual Natural Killer cells isolated in arrays of nanoliter wells2010Conference paper (Refereed)
  • 36.
    Guldevall, Karolin
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Gustafsson, Karin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Forslund, Elin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Frisk, Thomas
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Manneberg, Otto
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Olofsson, Per E.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Tauriainen, Johanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Stikvoort, Arwen
    Karolinska Institute.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Mattsson, Jonas
    Karolinska Institute.
    Kärre, Klas
    Karolinska Institute.
    Uhlin, Michael
    Karolinska Institute.
    Önfelt, Bjorn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Microchip screening platform for assessment of cytotoxic effector cellsManuscript (preprint) (Other academic)
    Abstract [en]

    Here we report a screening platform for assessment of the cytotoxic potential of individual natural killer (NK) or T cells within larger populations. Human primary NK cells or human Epstein-Barr virus (EBV)- specific T cells were distributed across a silicon-glass microchip containing 32 400 individual microwells loaded with target cells. Through fluorescence screening and automated image analysis the numbers of effector and live or dead target cells in each well could be assessed at different time-points after initial mixing. Cytotoxicity was also studied by time-lapse live-cell imaging in microwells quantifying the killing potential of individual NK cells. Although most resting NK cells (≈75%) were non-cytotoxic to the leukemia cell line K562, some NK cells were able to kill several (≥3) target cells within the 12 hours long experiment. We demonstrate that this assay can be used to enumerate and characterize cytotoxic cells, something that could find clinical applications, e.g. in the selection of donors for stem cell transplantation or generation of highly specific and cytotoxic cells for adoptive immunotherapy.

  • 37.
    Guldevall, Karolin
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Frisk, Thomas
    Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet.
    Hurtig, Johan
    Department of Chemsitry, University of Washington, Seattle, USA.
    Christakou, Athanasia
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Manneberg, Otto
    Department of Environmental Health, Harvard School of Public Health, Boston, USA.
    Lindström, Sara
    KTH, School of Biotechnology (BIO), Nano Biotechnology.
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO), Nano Biotechnology.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Imaging Immune Surveillance of Individual Natural Killer Cells Confined in Microwell Arrays2010In: PLOS ONE, ISSN 1932-6203, Vol. 5, no 11, p. e15453-Article in journal (Refereed)
    Abstract [en]

    New markers are constantly emerging that identify smaller and smaller subpopulations of immune cells. However, there is a growing awareness that even within very small populations, there is a marked functional heterogeneity and that measurements at the population level only gives an average estimate of the behaviour of that pool of cells. New techniques to analyze single immune cells over time are needed to overcome this limitation. For that purpose, we have designed and evaluated microwell array systems made from two materials, polydimethylsiloxane (PDMS) and silicon, for high-resolution imaging of individual natural killer (NK) cell responses. Both materials were suitable for short-term studies (<4 hours) but only silicon wells allowed long-term studies (several days). Time-lapse imaging of NK cell cytotoxicity in these microwell arrays revealed that roughly 30% of the target cells died much more rapidly than the rest upon NK cell encounter. This unexpected heterogeneity may reflect either separate mechanisms of killing or different killing efficiency by individual NK cells. Furthermore, we show that high-resolution imaging of inhibitory synapse formation, defined by clustering of MHC class I at the interface between NK and target cells, is possible in these microwells. We conclude that live cell imaging of NK-target cell interactions in multi-well microstructures are possible. The technique enables novel types of assays and allow data collection at a level of resolution not previously obtained. Furthermore, due to the large number of wells that can be simultaneously imaged, new statistical information is obtained that will lead to a better understanding of the function and regulation of the immune system at the single cell level.

  • 38.
    Hammer, Q.
    et al.
    Karolinska Inst, Dept Med Huddinge, Ctr Infect Med, Stockholm, Sweden..
    Perica, K.
    Mem Sloan Kettering Canc Ctr, 1275 York Ave, New York, NY 10021 USA..
    Mbofung, R. M.
    Fate Therapeut, San Diego, CA USA..
    van Ooijen, Hanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Goodridge, J. P.
    Fate Therapeut, San Diego, CA USA..
    Valamehr, B.
    Fate Therapeut, San Diego, CA USA..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Sadelain, M.
    Mem Sloan Kettering Canc Ctr, 1275 York Ave, New York, NY 10021 USA..
    Malmberg, K. J.
    Karolinska Inst, Dept Med Huddinge, Ctr Infect Med, Stockholm, Sweden.;Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Genetic ablation of adhesion ligands effectively averts rejection of allogeneic immune cells2022In: European Journal of Immunology, ISSN 0014-2980, E-ISSN 1521-4141, Vol. 52, p. 23-23Article in journal (Other academic)
  • 39.
    Hammer, Quirin
    et al.
    Karolinska Inst, Ctr Infect Med, Dept Med, Huddinge, Sweden..
    Perica, Karlo
    Mem Sloan Kettering Canc Ctr, New York, NY USA..
    Mbofung, Rina M.
    Fate Therapeut Inc, San Diego, CA USA..
    van Ooijen, Hanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Varady, Erika
    Fate Therapeut Inc, San Diego, CA USA..
    Jelcic, Mark
    Fate Therapeut Inc, San Diego, CA USA..
    Pan, Yijia
    Fate Therapeut Inc, San Diego, CA USA..
    Momayyezi, Pouria
    Karolinska Inst, Dept Med, Ctr Infect Med, Stockholm, Sweden..
    Groff, Brian
    Fate Therapeut Inc, San Diego, CA USA..
    Abujarour, Ramzey
    Fate Therapeut Inc, San Diego, CA USA..
    Krokeide, Silje Zandstra
    Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Lee, Tom
    Fate Therapeut Inc, San Diego, CA USA..
    Williams, Alan M.
    Fate Therapeut Inc, San Diego, CA USA..
    Goodridge, Jode P.
    Fate Therapeut Inc, San Diego, CA USA..
    Valamehr, Bahram
    Fate Therapeut Inc, San Diego, CA USA..
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Sadelain, Michel
    Mem Sloan Kettering Canc Ctr, Ctr Cell Engn, New York, NY USA..
    Malmberg, Karl-Johan
    Karolinska Inst, Dept Med, Ctr Infect Med, Stockholm, Sweden.;Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Combined Genetic Ablation of CD54 and CD58 in CAR Engineered Cytotoxic Lymphocytes Effectively Averts Allogeneic Immune Cell Rejection2022In: Blood, ISSN 0006-4971, E-ISSN 1528-0020, Vol. 140, no Supplement 1, p. 1165-1166Article in journal (Other academic)
    Abstract [en]

    Allogeneic cell therapies hold promise to be cost effective with scaled manufacturing for multi-dosing and on-demand off-the-shelf availability. A critical consideration for allogeneic cell products is their ability to persist, maintain function and avoid rejection by the patient's immune system. Genetic knockout (KO) of beta-2-microglobulin (B2M) leads to complete loss of cell-surface human leukocyte antigen (HLA) class I expression and efficiently abrogates CD8+ T-cell reactivity. However, loss of HLA class I triggers NK cell-mediated missing-self recognition and manipulation of B2M must therefore be combined with other immune-modulating strategies to limit recipient NK cell reactivity.We hypothesized that rejection by the patient's immune system can be diminished in primary CAR T cells, iPSC-derived T (iT) and NK (iNK) cells by reverse-engineering common tumor escape mechanisms. The adhesion molecules CD54 and CD58 are both present at the target cell side of the immune synapse, and loss of either of these molecules have previously been reported to elicit immune escape. Here, we show that the combined deletion of CD54 and CD58 in allogeneic immune effector cells makes them resistant to rejection by recipient immune cells through unidirectional reduced synapse formation (Figure 1A).HLA class I down-regulation by B2M silencing in primary T and NK cells triggered potent cytotoxicity by resting allogeneic NK cells. This response was mostly driven by educated NK cells expressing either NKG2A or killer cell immunoglobulin-like receptors (KIR) binding to HLA-E and HLA-C, respectively. However, over-expression of HLA-E or single HLA-C ligands in a K562 screening model only shut down the specific response of the NK cell subset carrying the cognate inhibitory receptor, resulting in only partial resistance to NK cells at the bulk level. Notably, the introduction of HLA-E was particularly detrimental in donors with expanded NKG2C+ NK cell subsets, due to its stimulatory effect through the activating NKG2C receptor. In contrast, combined deletion of CD54 and CD58 in target cells uniquely decreased the response of all tested NK cell subsets and showed universal reduction across NK cell populations from 18 healthy donors (Figure 1B). To delineate the mechanisms behind the increased resistance of target cells carrying these edits, we studied NK cell-target cell interactions at the single cell level by confocal microscopy in microchips. Allogeneic NK cells formed fewer conjugates and failed to form productive immune synapses with CD54-/-CD58-/- target cells, supporting the notion that they are more resistant to NK-cell mediated killing by unidirectional altered adhesion.We next introduced these edits in primary B2M-/- T cells engineered to express a second generation CAR19 from the TRAC locus. Corroborating the K562 screen, CD54-/-CD58-/-B2M-/- CAR-T cells had a selective survival advantage over B2M-/- CAR T cells and HLA-E-over-expressing B2M-/- CAR T cells in conventional mixed lymphocyte reaction (MLR) assays in vitro. Furthermore, we established an in vivo model to probe the effect of different genetic edits on the persistence of allogeneic cell therapy products. To this end, a mixed population of B2M-/- CAR T cells additionally bearing either CD54 and/or CD58 KO, HLA-E over-expression, or no further edits were infused into mice harboring allogeneic healthy donor PBMC. We found that CD54-/-CD58-/-B2M-/- CAR T cells had significantly better in vivo persistence compared to both B2M-/- CAR T cells and HLA-E+B2M-/- CAR T cells in the presence of PBMC from healthy donors (Figure 1B).Although multiplexed editing is feasible in primary CAR T cells, the iPSC platform has an unmatched capacity for homogenously introducing multiple immune-evasion strategies for off-the-shelf cell therapy. Similar to primary CAR T cells, multiplexed edited CD54-/-CD58-/-B2M-/-CIITA-/- iNK cells showed normal growth kinetics and were resistant to rejection by activated allogeneic NK cells in MLR assays.Together, these data demonstrate that reverse-engineering of common tumor escape mechanisms, which render target cells less susceptible to immune synapse formation, is an effective strategy to avert immune rejection of allogeneic CAR T and iPSC-derived CAR NK cells.

  • 40.
    Hammer, Quirin
    et al.
    Center for Infectious Medicine, Department of Medicine, Karolinska Institutet.
    Perica, Karlo
    Center for Cell Engineering, Memorial Sloan Kettering Cancer Center, New York, USA.
    van Ooijen, Hanna
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Mbofung, Rina
    Fate Therapeutics Inc. San Diego, US.
    Momayyezi, Pouria
    Center for Infectious Medicine, Department of Medicine, Karolinska Institutet.
    Varady, Erika
    Fate Therapeutics Inc. San Diego US.
    Martin, Karen E.
    Department of Cancer Immunology, Oslo Univeristy Hospital.
    Pan, Yijia
    Fate Therapeutics Inc. San diego, US.
    Jelcic, Mark
    Fate Therapeutics Inc. San Diego, US.
    Groff, Brian
    Fate Therapeutics Inc. San Diego, US.
    Abujarour, Ramzey
    Fate Therapeutics Inc. San Diego. US.
    Krokeide, Silje
    Department of Cancer Immunology, Oslo University Hospital.
    Lee, Tom
    Fate Therapeutics Inc. San Diego, US.
    Williams, Alan
    Fate Therapeutics Inc. San Diego, US.
    Goodridge, Jode P.
    Fate Therapeutics Inc. San Diego, US.
    Valamehr, Bahram
    Fate Therapeutics Inc. San Diego, US.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab.
    Sadelain, Michel
    Center for Cell Engineering, Memorial Sloan Kettering Cancer Center, New York, US.
    Malmberg, Karl-Johan
    Center for Infectious Medicine, Department of Medicine, Karolinska Institutet.
    Genetic ablation of adhesion ligands averts rejection of allogeneic immune cellsManuscript (preprint) (Other academic)
    Abstract [en]

    A significant barrier to the broad clinical implementation of allogeneic cell therapies is rejection of the allogeneic cells by the recipient immune system. Silencing of β-2-microglobulin (B2M) expression is a commonly employed strategy to evade T cell mediated rejection; however, the absence of B2M triggers missing-self responses by recipient natural killer (NK) cells. Here, we demonstrate that deletion of the adhesionligands CD54 and CD58 on allogeneic cells dampens recipient NK cell reactivityindependent of NK cell sub-population and inhibitory receptor expression. Additionally,genetic ablation of CD54 and CD58 in B2M-deficient allogeneic CAR T cells andinduced pluripotent stem cell-derived NK (iPSC-NK) cells reduces their susceptibility to rejection by recipient NK cells both in vitro and in vivo without affecting their anti-tumor effector potential. Thus, these data show that genetic ablation of adhesion ligands effectively mitigates rejection of allogeneic immune cells, enabling the generation of rejection-resistant allogeneic cell products.

  • 41.
    Haroun-Izquierdo, Alvaro
    et al.
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Vincenti, Marianna
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Netskar, Herman
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    van Ooijen, Hanna
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics.
    Zhang, Bin
    Univ Minnesota, Masonic Canc Ctr, Minneapolis, MN USA..
    Bendzick, Laura
    Univ Minnesota, Masonic Canc Ctr, Minneapolis, MN USA..
    Kanaya, Minoru
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Momayyezi, Pouria
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Li, Shuo
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Wiiger, Merete Thune
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Hoel, Hanna Julie
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Krokeide, Silje Zandstra
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Kremer, Veronika
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Tjonnfjord, Geir
    Univ Oslo, Oslo Univ Hosp, Inst Clin Med, Dept Hematol, Oslo, Norway.;Univ Oslo, Inst Clin Med, KG Jebsen Ctr B cell malignancies, Oslo, Norway..
    Berggren, Stephanie
    Karolinska Univ Hosp, Karolinska Ctr, Cell Therapy Clin Res Ctr, Vecura, Stockholm, Sweden..
    Wikström, Kristina
    Karolinska Univ Hosp, Karolinska Ctr, Cell Therapy Clin Res Ctr, Vecura, Stockholm, Sweden..
    Blomberg, Pontus
    Karolinska Univ Hosp, Karolinska Ctr, Cell Therapy Clin Res Ctr, Vecura, Stockholm, Sweden..
    Alici, Evren
    Karolinska Inst, Ctr Hematol & Regenerat Med, Dept Med Huddinge, Stockholm, Sweden..
    Felices, Martin
    Univ Minnesota, Masonic Canc Ctr, Minneapolis, MN USA..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Microbiol Tumour & Cell Biol, Stockholm, Sweden..
    Höglund, Petter
    Karolinska Inst, Ctr Hematol & Regenerat Med, Dept Med Huddinge, Stockholm, Sweden.;Karolinska Univ Hosp, Dept Clin Immunol & Transfus Med, Stockholm, Sweden..
    Valamehr, Bahram
    Fate Therapeut Inc, La Jolla, CA USA..
    Ljunggren, Hans-Gustaf
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Björklund, Andreas
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Hammer, Quirin
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Kveberg, Lise
    Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Cichocki, Frank
    Univ Minnesota, Masonic Canc Ctr, Minneapolis, MN USA..
    Miller, Jeffrey S.
    Univ Minnesota, Masonic Canc Ctr, Minneapolis, MN USA..
    Malmberg, Karl-Johan
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden.;Univ Oslo, Oslo Univ Hosp, Inst Canc Res, Dept Canc Immunol, Oslo, Norway..
    Sohlberg, Ebba
    Karolinska Inst, Ctr Infect Med, Dept Med Huddinge, Stockholm, Sweden..
    Adaptive single-KIR(+)NKG2C(+) NK cells expanded from select superdonors show potent missing-self reactivity and efficiently control HLA-mismatched acute myeloid leukemia2022In: Journal for ImmunoTherapy of Cancer, E-ISSN 2051-1426, Vol. 10, no 11, p. e005577-, article id e005577Article in journal (Refereed)
    Abstract [en]

    BackgroundNatural killer (NK) cells hold great promise as a source for allogeneic cell therapy against hematological malignancies, including acute myeloid leukemia (AML). Current treatments are hampered by variability in NK cell subset responses, a limitation which could be circumvented by specific expansion of highly potent single killer immunoglobulin-like receptor (KIR)(+)NKG2C(+) adaptive NK cells to maximize missing-self reactivity.MethodsWe developed a GMP-compliant protocol to expand adaptive NK cells from cryopreserved cells derived from select third-party superdonors, that is, donors harboring large adaptive NK cell subsets with desired KIR specificities at baseline. We studied the adaptive state of the cell product (ADAPT-NK) by flow cytometry and mass cytometry as well as cellular indexing of transcriptomes and epitopes by sequencing (CITE-Seq). We investigated the functional responses of ADAPT-NK cells against a wide range of tumor target cell lines and primary AML samples using flow cytometry and IncuCyte as well as in a mouse model of AML.ResultsADAPT-NK cells were >90% pure with a homogeneous expression of a single self-HLA specific KIR and expanded a median of 470-fold. The ADAPT-NK cells largely retained their adaptive transcriptional signature with activation of effector programs without signs of exhaustion. ADAPT-NK cells showed high degranulation capacity and efficient killing of HLA-C/KIR mismatched tumor cell lines as well as primary leukemic blasts from AML patients. Finally, the expanded adaptive NK cells had preserved robust antibody-dependent cellular cytotoxicity potential and combination of ADAPT-NK cells with an anti-CD16/IL-15/anti-CD33 tri-specific engager led to near-complete killing of resistant CD45(dim) blast subtypes.ConclusionsThese preclinical data demonstrate the feasibility of off-the-shelf therapy with a non-engineered, yet highly specific, NK cell population with full missing-self recognition capability.

  • 42. Hsu, Hsiang-Ting
    et al.
    Mace, Emily M.
    Carisey, Alexandre F.
    Viswanath, Dixita I.
    Christakou, Athanasia
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Wiklund, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Önfelt, Bjorn
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Orange, Jordan S.
    NK cells converge lytic granules to promote cytotoxicity and prevent bystander killing2016In: Journal of Cell Biology, ISSN 0021-9525, E-ISSN 1540-8140, Vol. 215, no 6, p. 875-889Article in journal (Refereed)
    Abstract [en]

    Natural killer (NK) cell activation triggers sequential cellular events leading to destruction of diseased cells. We previously identified lytic granule convergence, a dynein-and integrin signal-dependent movement of lysosome-related organelles to the microtubule-organizing center, as an early step in the cell biological process underlying NK cell cytotoxicity. Why lytic granules converge during NK cell cytotoxicity, however, remains unclear. We experimentally controlled the availability of human ligands to regulate NK cell signaling and promote granule convergence with either directed or nondirected degranulation. By the use of acoustic trap microscopy, we generated specific effector-target cell arrangements to define the impact of the two modes of degranulation. NK cells with converged granules had greater targeted and less nonspecific "bystander" killing. Additionally, NK cells in which dynein was inhibited or integrin blocked under physiological conditions demonstrated increased nondirected degranulation and bystander killing. Thus, NK cells converge lytic granules and thereby improve the efficiency of targeted killing and prevent collateral damage to neighboring healthy cells.

  • 43. Hurtig, Johan
    et al.
    Chiu, Daniel T.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Intercellular nanotubes: insights from imaging studies and beyond2010In: WILEY INTERDISCIPLINARY REVIEWS-NANOMEDICINE AND NANOBIOTECHNOLOGY, ISSN 1939-5116, Vol. 2, no 3, p. 260-276Article, review/survey (Refereed)
    Abstract [en]

    Cell-cell communication is critical to the development, maintenance, and function of multicellular organisms. Classical mechanisms for intercellular communication include secretion of molecules into the extracellular space and transport of small molecules through gap junctions. Recent reports suggest that cells also can communicate over long distances via a network of transient intercellular nanotubes. Such nanotubes have been shown to mediate intercellular transfer of organelles as well as membrane components and cytoplasmic molecules. Moreover, intercellular nanotubes have been observed in vivo and have been shown to enhance the transmission of pathogens such as human immunodeficiency virus (HIV)-1 and prions in vitro. These studies indicate that intercellular nanotubes may play a role both in normal physiology and in disease. (C) 2010 John Wiley & Sons, Inc. WIREs Nanomed Nanobiotechnol 2010 2 260-276

  • 44.
    Kamali-Zare, Padideh
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Kowalewski, Jacob M
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Zelenina, Marina
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Aperia, Anita
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Role of diffusion limited space on water and salt homeostasisManuscript (preprint) (Other academic)
  • 45.
    Kamali-Zare, Padideh
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Kowalewski, Jacob
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Zelenina, Marina
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Aperia, Anita
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Diffusion limited space contributes to K+ siphoning by regulation of K+ and water homeostasis in astrocytesManuscript (preprint) (Other academic)
    Abstract [en]

    Diffusion Limited Space (DLS) is defined as a region where diffusion is limited by the geometry. Two examples of DLS in the brain are the neuronal synapse, and the narrow region between astrocyte endfeet and blood capillaries. In a series of geometrical models we show that DLS plays a role in regulation of water and K+ homeostasis in the brain by an indirect functional coupling of aquaporins (AQPs) and inward rectifying K+ (Kir) channels in a membrane microdomain.

    1. Simulations in geometrical models of a synapse region show that following a step increase in synaptic [K+], both K+ and water are taken up by astrocytes via AQPs and Kir channels lining the synapse.  This uptake creates a transient depletion of water in the synapse region that, enhanced by the DLS, facilitates K+ uptake and an efficient clearance of excess K+ from the synapse.

    2. Simulations in a geometrical model of astrocytes show that the DLS formed between astrocyte endfeet and blood capillaries, facilitate the siphoning of accumulated K+ into the extracellular space facing the blood capillaries. The DLS geometry creates an efficient coupling between AQPs and Kir channels.

    3. Furthermore, the models show that a local coupling between water and K+ transport is important for the maintenance of membrane potential and the net K+ spatial buffering capacity in the astrocytes.

    4. In the full geometrical model of K+ spatial buffering we show that the geometry of the extracellular space both in the synapse region and in the endfeet is an essential component for the cell volume regulation.

    Our results suggest that for regulation of K+ and water homeostasis in astrocytes, not only the classical aspects of functional couplings between proteins, but also the geometry of the cell and the microdomains are important. Further, our results suggest a central role for AQPs in the astrocyte endfeet and identify their contribution to K+ siphoning.

  • 46.
    Karampatzakis, Alexandros
    et al.
    Univ Manchester, Fac Biol Med & Hlth, Lydia Becker Inst Immunol, Manchester, Lancs, England..
    Broz, Petr
    Univ Manchester, Fac Biol Med & Hlth, Lydia Becker Inst Immunol, Manchester, Lancs, England..
    Rey, Camille
    Univ Manchester, Fac Biol Med & Hlth, Lydia Becker Inst Immunol, Manchester, Lancs, England..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics. KTH, Centres, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Microbiol Tumor & Cell Biol, Stockholm, Sweden..
    Cruz De Matos, Gabriela Dos Santos
    GlaxoSmithKline, Stevenage, Herts, England..
    Rycroft, Daniel
    GlaxoSmithKline, Stevenage, Herts, England..
    Ambrose, Ashley
    Univ Manchester, Fac Biol Med & Hlth, Lydia Becker Inst Immunol, Manchester, Lancs, England..
    Davis, Daniel M.
    Univ Manchester, Fac Biol Med & Hlth, Lydia Becker Inst Immunol, Manchester, Lancs, England..
    Antibody Afucosylation Augments CD16-Mediated Serial Killing and IFN gamma Secretion by Human Natural Killer Cells2021In: Frontiers in Immunology, E-ISSN 1664-3224, Vol. 12, article id 641521Article in journal (Refereed)
    Abstract [en]

    One mechanism by which monoclonal antibodies (mAb) help treat cancer or autoimmune disease is through triggering antibody-dependent cellular cytotoxicity (ADCC) via CD16 on Natural Killer (NK) cells. Afucosylation is known to increase the affinity of mAbs for CD16 on NK cells and here, we set out to assess how mAb afucosylation affects the dynamics of NK cell interactions, receptor expression and effector functions. An IgG1 version of a clinically important anti-CD20 mAb was compared to its afucosylated counterpart (anti-CD20-AF). Opsonization of CD20-expressing target cells, 721.221 or Daudi, with anti-CD20-AF increased NK cell cytotoxicity and IFN gamma secretion, compared to anti-CD20. The afucosylated mAb also caused a more rapid and greater loss of CD16 from NK cell surfaces. Loss of CD16 has recently been shown to be important for NK cell detachment and sequential engagement of multiple target cells. Here, live-cell time-lapse microscopy of individual cell-cell interactions in an aqueous environment and a three-dimensional matrix, revealed that anti-CD20-AF induced more rapid killing of opsonized target cells. In addition, NK cells detached more quickly from target cells opsonized with anti-CD20-AF compared to anti-CD20, which increased engagement of multiple targets and enabled a greater proportion of NK cells to perform serial killing. Inhibition of CD16 shedding with TAPI-0 led to reduced detachment and serial killing. Thus, disassembly of the immune synapse caused by loss of cell surface CD16 is a factor determining the efficiency of ADCC and antibody afucosylation alters the dynamics of intercellular interactions to boost serial killing.

  • 47.
    Khorshidi, Mohammad A.
    et al.
    KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Natural Killer Cell-Mediated Tumor Surveillance: Correlation Between Killing Efficiency, Transient Migration Behavior and Morphology2012In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 102, no 3, p. 706A-706AArticle in journal (Other academic)
  • 48.
    Khorshidi, Mohammad Ali
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Vanherberghen, Bruno
    KTH, School of Engineering Sciences (SCI), Applied Physics.
    Kowalewski, Jacob M.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Garrod, Kym R.
    Lindström, Sara
    KTH, School of Biotechnology (BIO), Nano Biotechnology.
    Andersson-Svahn, Helene
    KTH, School of Biotechnology (BIO), Nano Biotechnology.
    Brismar, Hjalmar
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Cahalan, Michael D.
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics.
    Analysis of transient migration behavior of natural killer cells imaged in situ and in vitro2011In: Integrative Biology, ISSN 1757-9694, E-ISSN 1757-9708, Vol. 3, no 7, p. 770-778Article in journal (Refereed)
    Abstract [en]

    We present a simple method for rapid and automatic characterization of lymphocyte migration from time-lapse fluorescence microscopy data. Time-lapse imaging of natural killer (NK) cells in vitro and in situ, both showed that individual cells transiently alter their migration behavior. Typically, NK cells showed periods of high motility, interrupted by transient periods of slow migration or almost complete arrests. Analysis of in vitro data showed that these periods frequently coincided with contacts with target cells, sometimes leading to target cell lysis. However, NK cells were also commonly observed to stop independently of contact with other cells. In order to objectively characterize the migration of NK cells, we implemented a simple method to discriminate when NK cells stop or have low motilities, have periods of directed migration or undergo random movement. This was achieved using a sliding window approach and evaluating the mean squared displacement (MSD) to assess the migration coefficient and MSD curvature along trajectories from individual NK cells over time. The method presented here can be used to quickly and quantitatively assess the dynamics of individual cells as well as heterogeneity within ensembles. Furthermore, it may also be used as a tool to automatically detect transient stops due to the formation of immune synapses, cell division or cell death. We show that this could be particularly useful for analysis of in situ time-lapse fluorescence imaging data where most cells, as well as the extracellular matrix, are usually unlabelled and thus invisible.

  • 49.
    Kienzle, T.
    et al.
    Karolinska Inst, Stockholm, Sweden..
    Zambarda, Chiara
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Nasi, A.
    Swedish StromaBio, Solna, Sweden..
    Önfelt, Björn
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biophysics.
    Kadri, N.
    Karolinska Inst, Stockholm, Sweden..
    Characterization of interactions between monocytes and mesenchymal stromal cells2023In: Cytotherapy, ISSN 1465-3249, E-ISSN 1477-2566, Vol. 25, no 6, p. S63-S63Article in journal (Other academic)
  • 50.
    Kördel, Mikael
    et al.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Fogelqvist, Emelie
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Carannante, Valentina
    Department of Microbiology, Karolinska Institutet.
    Önfelt, Björn
    KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Reddy, Hemanth K. N.
    Department of Cell and Molecular Biology, Uppsala University.
    Okamoto, Kenta
    Department of Cell and Molecular Biology, Uppsala University.
    Svenda, Martin
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Sellberg, Jonas A.
    KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.
    Hertz, Hans
    KTH, School of Engineering Sciences (SCI), Physics. KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics. KTH, School of Biotechnology (BIO), Centres, Albanova VinnExcellence Center for Protein Technology, ProNova.
    Biological Laboratory X-ray Microscopy2018In: Microscopy and Microanalysis, ISSN 1431-9276, E-ISSN 1435-8115, Vol. 24, no S2, p. 346-347Article in journal (Refereed)
123 1 - 50 of 118
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf