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Analysis of intracellular regulatory proteins by immunoaffinity capillary electrophoresis coupled with laser‐induced fluorescence detection
Author(s) -
Phillips Terry M.,
Smith Paul
Publication year - 2003
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.240
Subject(s) - capillary electrophoresis , chemistry , intracellular , analyte , antibody , microbiology and biotechnology , fluorescence , chromatography , biochemistry , biology , immunology , physics , quantum mechanics
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. Copyright © 2003 John Wiley & Sons, Ltd.