LASER BASED CELL LOADING & LYSIS: SINGLE CELL ELECTROPHORESIS: KINASE ACTIVITY

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Significant progress has been made toward specific aim 3. Currently, single adherent cells are lysed in less than 33 msec by a laser-induced shock wave. The contents of single cells are immediately loaded into a capillary, separated by electrophoresis within the capillary, and detected by laser-induced fluorescence. This was demonstrated and published in 1998 in Analytical Chemistry, vol 70, number 21, pages 4570-4577. Preliminary results show that kinase-substrate peptides can be measured from single mammalian cells. Further, cellular kinase activity can be measured as changes in the degree of substrate phosphorylation. Experiments are in progress to define parameters for and demonstrate physiologically relevant measurements. Specific aims 1 and 2 are yet to be addressed substantively, but will be accelerated by the completion of the current studies on adherent cells. Specific aim 4 will become possible immediately thereafter.