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Interfacing picoliter droplet microfluidics with addressable microliter compartments using fluorescence activated cell sorting
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.ORCID-id: 0000-0001-5232-0805
KTH, Skolan för bioteknologi (BIO), Proteomik och nanobioteknologi. KTH, Centra, Science for Life Laboratory, SciLifeLab.
2014 (Engelska)Ingår i: Sensors and actuators. B, Chemical, ISSN 0925-4005, E-ISSN 1873-3077, Vol. 194, s. 249-254Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Droplet microfluidic platforms have, while enabling high-throughput manipulations and the assaying of single cell scale compartments, been lacking interfacing to allow macro scale access to the output from droplet microfluidic operations. Here, we present a simple and high-throughput method for individually directing cell containing droplets to an addressable and macro scale accessible microwell slide for downstream analysis. Picoliter aqueous droplets containing low gelling point agarose and eGFP expressing Escherichia coli (E. coli) are created in a microfluidic device, solidified to agarose beads and transferred into an aqueous buffer. A Fluorescence activated cell sorter (FACS) is used to sort agarose beads containing cells into microwells in which the growth and expansion of cell colonies is monitored. We demonstrate fast sorting and high accuracy positioning of sorted 15 μm gelled droplet agarose beads into microwells (14 × 48) on a 25 mm × 75 mm microscope slide format using a FACS with a 100 μm nozzle and an xy-stage. The interfacing method presented here enables the products of high-throughput or single cell scale droplet microfluidics assays to be output to a wide range of microtiter plate formats familiar to biological researchers lowering the barriers for utilization of these microfluidic platforms.

Ort, förlag, år, upplaga, sidor
2014. Vol. 194, s. 249-254
Nyckelord [en]
Droplet microfluidics, Fluorescence activated cell sorting, Agarose beads, Microwell slide, Escherichia coli
Nationell ämneskategori
Biomedicinsk laboratorievetenskap/teknologi
Identifikatorer
URN: urn:nbn:se:kth:diva-140109DOI: 10.1016/j.snb.2013.12.089ISI: 000331575400033Scopus ID: 2-s2.0-84892492768OAI: oai:DiVA.org:kth-140109DiVA, id: diva2:688578
Forskningsfinansiär
Science for Life Laboratory - a national resource center for high-throughput molecular bioscienceVetenskapsrådet
Anmärkning

QC 20140117

Tillgänglig från: 2014-01-17 Skapad: 2014-01-17 Senast uppdaterad: 2017-12-06Bibliografiskt granskad
Ingår i avhandling
1. Miniaturised Microwell-based Cell Assays
Öppna denna publikation i ny flik eller fönster >>Miniaturised Microwell-based Cell Assays
2014 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

Cell heterogeneity in genetically identical cell populations is becoming a well-known and important phenomenon in cell biology. Current methods commonly utilise population-based analysis founded on averaged result. Hence there is a need for high-throughput cell assays on the single-cell level. By using miniaturised devices it is possible to enhance spatial and temporal control of the individual cells, increase the potential throughput and minimise the needed sample and reagent volume while enabling a wide range of biological applications.

This thesis is based on the results generated with a miniaturised microwell slide for cell assays. The microwell slide’s high-throughput compartmentalised configuration enables several hundred isolated experiments to be run simultaneously. The bottom of the wells is made out of a thin glass slide, which supports high-resolution imaging. The slide has a standardised format and its’ compatibility with conventional instruments is used extensively throughout the thesis. The presented papers all contribute to the development of the microwell slide by adding technical value or increasing the number of potential applications. For example, the slide was success-fully implemented as a chip-to-world output format for single microfluidic droplets in Paper I, by interfacing two miniaturised systems with fluorescence-activated cell sorting. In Paper II and III, microfluidic channels were integrated to increase spatial and temporal control of the added samples and reagents. In Paper II an automated stepwise concentration gradient generator was developed delivering a drug gradient to adherent mammalian cells in designated wells. In Paper III fluidic-imposed shear stress on endothelial cells was studied. In Paper IV, the slide was functionalised by coating the surfaces of the wells with several antibiotics at a defined concentration range. The coated slide was used for multiplex antibiotic susceptibility testing of bacterial pathogens, using an algorithm-based identification of the point defining lag to exponential phase transition. It successfully determined the pathogens susceptibility profile in 3-5 hours. Finally, in Paper V, a method to retrieve bacteria colonies with a desired phenotype from the wells for downstream genetic analysis was developed. In summary, the presented work has furthered the development of miniaturised high-throughput tools for various cell heterogeneity assays.

Ort, förlag, år, upplaga, sidor
Stockholm: KTH Royal Institute of Technology, 2014. s. iv, 70
Serie
TRITA-BIO-Report, ISSN 1654-2312 ; 2014:2
Nyckelord
microwell, miniaturisation, high-resolution imaging, high-throughput, cell culture, single-cell, clone, heterogeneity, antibiotic susceptibility testing, concentration gradient, interfacing, microchannels, cell retrieval
Nationell ämneskategori
Biomedicinsk laboratorievetenskap/teknologi
Identifikatorer
urn:nbn:se:kth:diva-140113 (URN)978-91-7501-982-6 (ISBN)
Disputation
2014-02-07, Air & Fire - Gamma building, Tomtebodavägen 23A, Solna, 10:00 (Engelska)
Opponent
Handledare
Anmärkning

QC 20140117. The abstract published on January 24th, 2014.

Tillgänglig från: 2014-01-17 Skapad: 2014-01-17 Senast uppdaterad: 2014-01-24Bibliografiskt granskad

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Jönsson, Håkan

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Bai, YunpengWeibull, EmilieJönsson, HåkanAndersson Svahn, Helene
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Proteomik och nanobioteknologiScience for Life Laboratory, SciLifeLab
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