Analysis of intracellular regulatory proteins by immunoaffinity capillary electrophoresis coupled with laser-induced...

Measurement of intracellular regulatory proteins is of great importance in many areas of biomedical research. In this communication we describe an antibody-based capillary electrophoresis system equipped with an on-line laser-induced fluorescence detector capable of measuring intracellular proteins in cultures as low as 100 cells. The system demonstrated a high degree of precision and accuracy, being capable of detecting the fluorochrome-labeled analytes of interest at concentration of approximately 0.5 pg. We have used this instrument to study concentrations of the intracellular regulatory proteins STAT-1 and STAT-3, following stimulation of lymphocyte cultures with the inflammatory cytokine, IL-6. Using a combination of four antibodies specific for either STAT-1 or STAT-3 in both their nonphosphorylated and phosphorylated forms, we were able to demonstrate the differential expression of these proteins over time.