Reversibly sealed multilayer microfluidic device for integrated cell perfusion and on-line chemical analysis of cultured adipocyte secretions.
A three-layer microfluidic device was developed that combined perfusion of cultured cells with on-line chemical analysis for near real-time monitoring of cellular secretions. Two layers were reversibly sealed to form a cell chamber that allowed cells grown on coverslips to be loaded directly into the chip. The outlet of the chamber was in fluidic contact with a third layer that was permanently bonded. Perfusate from the cell chamber flowed into this third layer where a fluorescence enzyme assay for non-esterified fatty acid (NEFA) was performed on-line. The device was used to monitor efflux of NEFAs from approximately 6,200 cultured adipocytes with 83 s temporal resolution. Perfusion of murine 3T3-L1 cultured adipocytes resulted in an average basal concentration of 24.2 +/- 2.4 microM NEFA (SEM, n = 6) detected in the effluent corresponding to 3.31 x 10(-5) nmol cell(-1) min(-1). Upon pharmacological treatment with a beta-adrenergic agonist to stimulate lipolysis, a 6.9 +/- 0.7-fold (SEM, n = 6) sustained increase in NEFA secretion was observed. This multilayer device provides a versatile platform that could be adapted for use with other cell types to study corresponding cellular secretions in near real-time.
Clark
Anna M
AM
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
Sousa
Kyle M
KM
Chisolm
Claire N
CN
MacDougald
Ormond A
OA
Kennedy
Robert T
RT
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D018929:Cell Culture Techniques; D005230:Fatty Acids, Nonesterified / Q000032:analysis* / Q000378:metabolism; D008066:Lipolysis; D051379:Mice; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D009862:Online Systems / Q000295:instrumentation
D005230:Fatty Acids, Nonesterified