Mating cell traps to silica microbands for picoliter thin layer chromatography (pTLC) assay of phosphoinositide...

The phosphoinositide pathway plays key roles in the survival and proliferation of single tumor cells driving a need for single-cell assays of phosphoinositide signaling. In this work, cell traps aligned over monolithic silica microbands [picoliter thin layer chromatography (pTLC)] were used to place a single cell on microbands within an array with a greater than 70% efficiency. Separation and quantification of the metabolites of BODIPY fluorescein-labelled phosphoinositide 4-phosphate (BODIPY FL PI4P) formed was demonstrated in over 900 single cells using pTLC with cell traps. The major metabolite of BODIPY FL PI4P formed in cells was BODIPY FL phosphatidylinositol (PI) likely due to the actions of the suppressor of actin (SAC) family of phosphoinositide phosphatases. A minor metabolite possessed a migration time similar to that of BODIPY fluorescein-labelled phosphatidylinositol 4,5-bisphosphate (BODIPY FL PI(4,5)P₂). On average 59% ± 1% of BODIPY FL PI4P was dephosphorylated in K562 (chronic myelogenous leukemia), while significantly greater dephosphorylation (62% ± 1%) was observed in MM1S (multiple myeloma). In contrast the aggressive cell Kasumi-3 (undifferentiated acute myeloblastic leukemia) displayed significantly greater metabolism (66% ± 2%) of the PI4P reporter than other cell lines. All cell lines assessed displayed large numbers of cells with metabolic behavior far removed from that of the population average, revealing the full extent of cell-based behaviors in the cell population. Immunoassay of trapped Kasumi-3 cells followed by pTLC demonstrated the potential for pairing other assays with pTLC.