Microfluidic concentration-enhanced cellular kinase activity assay.
In this paper, we reported a simple, disposable PDMS micro/nanofluidic preconcentration chip for in vitro concentration-enhanced cell kinase assays. Utilizing the preconcentration (electrokinetic trapping) directly from cell lysate (1 mM ATP) samples, we could achieve at least a 25-fold increase in reaction velocity and 65-fold enhancement in sensitivity. In addition, we shorten the assay time down to less than 10 min, with the sample volume requirements of down to approximately 5 cells. This device could be a generic and powerful tool for diagnostics and systems biology studies at the single-cell level, if properly optimized and integrated with the cell culture microdevices.
Lee
Jeong Hoon
JH
Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
Cosgrove
Benjamin D
BD
Lauffenburger
Douglas A
DA
Han
Jongyoon
J
D053000:Analytic Sample Preparation Methods; D002457:Cell Extracts / Q000737:chemistry; D045744:Cell Line, Tumor; D002477:Cells / Q000201:enzymology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000378:metabolism*; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D036103:Nanotechnology / Q000295:instrumentation / Q000379:methods; D017346:Protein Serine-Threonine Kinases / Q000032:analysis / Q000378:metabolism*; D000099190:MAP-Kinase-Activated Kinase 2
D002457:Cell Extracts; D017868:Cyclic AMP-Dependent Protein Kinases; D047908:Intracellular Signaling Peptides and Proteins; D017346:Protein Serine-Threonine Kinases; D000099190:MAP-Kinase-Activated Kinase 2