Laser-micropipet combination for single-cell analysis.
Due to its potential for exquisite mass detection limits and resolving power, capillary electrophoresis is used for biochemical measurements on single cells; however, accurate measurements of many physiological parameters require sampling strategies that are considerably faster than those presently available. We have developed a laser-based technique to lyse single, adherent, mammalian cells on millisecond time scales. The cellular contents are then introduced into a capillary where electrophoretic separation and detection are performed. Improved temporal resolution of biological measurements results from the extremely rapid lysis made possible by this method. Additionally, the cell is not perturbed by mechanical or electrical stresses prior to sampling. Such disturbances can alter cellular physiology, resulting in inaccurate measurements. The fast cell lysis, the absence of cellular stresses prior to lysis, and the application to adherent mammalian cells are significant refinements to CE-based measurements on single cells. With this laser-micropipet combination, it will be possible to measure the intracellular concentration of molecules that change on subsecond to second time scales, for example, substrates of many cellular enzymes.
Sims
C E
CE
Department of Physiology and Biophysics, University of California, Irvine 92697, USA.
Meredith
G D
GD
Krasieva
T B
TB
Berns
M W
MW
Tromberg
B J
BJ
Allbritton
N L
NL
D000818:Animals; D002477:Cells / Q000737:chemistry*; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D005452:Fluoresceins; D007834:Lasers; D051381:Rats; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014407:Tumor Cells, Cultured
D005452:Fluoresceins; C018506:diacetylfluorescein