title: Microfluidic perfusion system for culturing and imaging yeast cell microarrays and rapidly exchanging media.
abstract: High resolution live cell microscopy is increasingly used to detect cellular dynamics in response to drugs and chemicals, but it depends on complex and expensive liquid handling devices that have limited its wider adoption. Here, we present a microfluidic perfusion system that is built without using specialized microfabrication infrastructure, simple to use because only a pipette is needed for liquid handling, and yet allows for rapid media exchange and simultaneous fluorescence microscopy imaging. Yeast cells may be introduced from a culture, or spotted as arrays on a coverslip, and are sandwiched with a 20 mum thick track-etched membrane. A second coverslip and a mesh with 120 mum porosity are placed on top, forming a microfluidic conduit for lateral flow of solutions by capillary effects. Solutions introduced through the inlet flow through the mesh and chemicals diffuse vertically across the membrane to the cells trapped below. Solutions are exchanged by adding a new sample to the inlet. Using this system, we studied the dynamic response of F-actin in living yeast expressing Sac6-EGFP-a protein associated with discrete F-actin structures called "patches"-to the drug latrunculin A, a well known inhibitor of actin polymerization. We observed that the patches disappeared in 85% of the cells within 5 min, and re-assembled in 45 min following exchange of the drug with media. The perfusion system presented here is a simple, inexpensive device suited for analysis of drug dose-response and regeneration of single cells and arrays of cells.
authors: Biomedical Engineering Department, McGill University, Montréal, QC, Canada.
authors: Maryam
authors: M
authors: Mirzaei
authors: Mateu
authors: M
authors: Pla-Roca
authors: Roozbeh
authors: R
authors: Safavieh
authors: Elena
authors: E
authors: Nazarova
authors: Mohammadali
authors: M
authors: Safavieh
authors: Huiyan
authors: H
authors: Li
authors: Jackie
authors: J
authors: Vogel
authors: David
authors: D
authors: Juncker
mesh_terms: D000199:Actins / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000295:instrumentation*; D003470:Culture Media; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D004867:Equipment Design; D046228:Microarray Analysis / Q000295:instrumentation*; D046210:Microfluidic Analytical Techniques; D008856:Microscopy, Fluorescence; D055616:Microtechnology; D057054:Molecular Imaging / Q000295:instrumentation*; D010477:Perfusion / Q000295:instrumentation*; D004718:Saccharomycetales / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D053778:Thiazolidines / Q000494:pharmacology; D013997:Time Factors
chemical_list: D000199:Actins; D019086:Bridged Bicyclo Compounds, Heterocyclic; D003470:Culture Media; D053778:Thiazolidines; C037067:latrunculin A