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Design and evaluation of a microfluidic system for inhibition studies of yeast cell signaling

Paper i proceeding
Författare Charlotte Hamngren Blomqvist
Peter Dinér
Morten Grøtli
Mattias Goksör
Caroline B. Adiels
Publicerad i Proceedings of SPIE - The International Society for Optical Engineering. Optical Trapping and Optical Micromanipulation IX, San Diego, 12-16 August 2012
Volym 8458
ISBN 978-08-19-49175-6
ISSN 0277-786X
Publiceringsår 2012
Publicerad vid Institutionen för kemi och molekylärbiologi
Institutionen för fysik (GU)
Språk en
Länkar dx.doi.org/10.1117/12.929728
https://gup.ub.gu.se/file/200666
Ämnesord Microfluidics, Optical manipulation, Signal transduction pathways, Single-cell analysis
Ämneskategorier Fysik

Sammanfattning

In cell signaling, different perturbations lead to different responses and using traditional biological techniques that result in averaged data may obscure important cell-to-cell variations. The aim of this study was to develop and evaluate a four-inlet microfluidic system that enables single-cell analysis by investigating the effect on Hog1 localization post a selective Hog1 inhibitor treatment during osmotic stress. Optical tweezers was used to position yeast cells in an array of desired size and density inside the microfluidic system. By changing the flow rates through the inlet channels, controlled and rapid introduction of two different perturbations over the cell array was enabled. The placement of the cells was determined by diffusion rates flow simulations. The system was evaluated by monitoring the subcellular localization of a fluorescently tagged kinase of the yeast "High Osmolarity Glycerol" (HOG) pathway, Hog1-GFP. By sequential treatment of the yeast cells with a selective Hog1 kinase inhibitor and sorbitol, the subcellular localization of Hog1-GFP was analysed on a single-cell level. The results showed impaired Hog1-GFP nuclear localization, providing evidence of a congenial design. The setup made it possible to remove and add an agent within 2 seconds, which is valuable for investigating the dynamic signal transduction pathways and cannot be done using traditional methods. We are confident that the features of the four-inlet microfluidic system will be a valuable tool and hence contribute significantly to unravel the mechanisms of the HOG pathway and similar dynamic signal transduction pathways.

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