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Microfluidic Centrifugation Assisted Precipitation based DNA Quantification
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.ORCID iD: 0000-0001-9869-7181
Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institute, Sweden..ORCID iD: 0000-0001-7398-8927
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology.
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Nano Biotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab. Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institute, Sweden..
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Nucleic acid amplification methods are increasingly being used to detect trace quantities of DNA in samples for various diagnostic applications. However, quantifying the amount of DNA from such methods often require time consuming purification, washing or labeling step. Here, we report a novel microfluidic centrifugation assisted precipitation (uCAP) method for single-step DNA quantification. The method is based on formation of a visible precipitate, that can be quantified, when an intercalating dye (GelRed) is added to DNA sample and centrifuged for few seconds. We describe the mechanism leading to the precipitation phenomenon. We utilize centrifugal microfluidics to precisely control the formation of visible and quantifiable mass. Using a standard CMOS sensor for imaging, we report a detection limit of 45 ng/ul. Furthermore, using an integrated Lab-on-DVD platform we recently developed, the detection limit was lowered to 10 ng/ul, which is comparable to current commercially available instruments for DNA quantification. As a proof of principle, we demonstrate the quantification of LAMP products for a HIV-1B type genome containing plasmid on the Lab-on-DVD platform. The simple DNA quantification system could facilitate advanced molecular diagnosis at point of care.

National Category
Medical and Health Sciences
Research subject
Biotechnology
Identifiers
URN: urn:nbn:se:kth:diva-244820OAI: oai:DiVA.org:kth-244820DiVA, id: diva2:1292345
Note

QC 20190228

Available from: 2019-02-27 Created: 2019-02-27 Last updated: 2019-03-12Bibliographically approved
In thesis
1. Point of care microfluidic tool development for resource limited settings
Open this publication in new window or tab >>Point of care microfluidic tool development for resource limited settings
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The development of point of care diagnostics using recent advances in microfluidics have the potential to transform health care in several ways, especially in resource limited settings with limited access to advanced health care infrastructure. However, translating a point of care device to reality is often a challenging task because of the complexities involved in integrating a number of diverse engineering concepts into an easy to use, accurate and portable device. This thesis focuses on miniaturization of crucial diagnostic laboratory tools, that can be used in a portable point of care format without compromising on the accuracy or performance. The first part of the thesis (Paper I-III) focuses on understanding and applying elasto-inertial microfluidics, which is a label-free and passive bio-particle sorting and separation method. A basic understanding of particle trajectories in both inertial (Paper I) and visco-elastic flows (Paper II) is established, followed by an investigation on the combined effects of inertia and elasticity (Paper III). The second part of the thesis (Paper IV-VI) focuses on developing integrated microfluidic platforms, each of which addresses different aspects of point of care diagnostic applications. The applications include neonatal diagnostics using a hand-driven Slipdisc technique (Paper IV), rapid nucleic acid quantification using a novel precipitate-based detection on a centrifugal microfluidics platform (Paper V), and hematocrit level measurement in blood using a portable lab-on- Disc platform operated by a mobile phone (Paper VI). The proof of concept microfluidic tools presented in the scope of this thesis have the potential to replace a number of functions of standard laboratory equipment, at a fraction of the price and without compromising performance. Hence, the different methods developed should contribute towards decentralization of medical testing laboratories, making healthcare accessible to one and all.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2019. p. 63
Series
TRITA-CBH-FOU ; 2019:15
Keywords
Blood, control, cell separation, centrifugal microfluidics, diagnostics, elasto-inertial, hematocrit level, microfluidics, neonatal diagnostics, nucleic acid quantification, point of care, particle focusing, resource limited settings.
National Category
Medical and Health Sciences Engineering and Technology
Research subject
Biotechnology
Identifiers
urn:nbn:se:kth:diva-244825 (URN)978-91-7873-122-0 (ISBN)
Public defence
2019-03-29, Air & Fire auditorium, Science for Life Laboratory, Tomtebodavägen 23, Solna, 10:00 (English)
Opponent
Supervisors
Note

QC 20190228

Available from: 2019-02-28 Created: 2019-02-28 Last updated: 2019-02-28Bibliographically approved

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Lapins, NoaZhang, WangKazemzadeh, AminRussom, Aman

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