Noncompetitive immunoassays using protein G affinity capillary chromatography and capillary electrophoresis with laser-induced fluorescence detection.
A new and simple approach to perform immunoassay using protein G affinity capillary chromatography and laser-induced fluorescence detection was described. A noncompetitive assay for monoclonal anti-bovine serum albumin (BSA) was used to test the performance of the system. Fluorescein isothiocyanate labeled BSA was used as a tracer to determine anti-BSA in pM level. Capillaries with inner diameter of 150 microns were packed with recombinant protein G-bound perfusive support. The packed capillary was used to capture the immunocomplexes, which were subsequently desorbed by 100 mM glycine (pH 9.0). Open tube capillary electrophoresis-based immunoassay (CEIA) for anti-BSA was also performed. Using standard samples, calibration curves for anti-BSA was established in both assays. Compared with CEIA, this system improved the concentration sensitivity for about 100-fold due to the pre-concentration of immunocomplex in the protein G column, while the mass sensitivity was similar in the two methods.
Wang
Q
Q
Department of Chemistry, Tsinghua University, Beijing, China.
Luo
G
G
Ou
J
J
Yeung
W S
WS
D000911:Antibodies, Monoclonal / Q000302:isolation & purification; D002846:Chromatography, Affinity / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D007118:Immunoassay / Q000379:methods*; D007834:Lasers; D009419:Nerve Tissue Proteins / Q000737:chemistry; D012710:Serum Albumin, Bovine / Q000276:immunology; D013050:Spectrometry, Fluorescence
D000911:Antibodies, Monoclonal; C029152:G-substrate; D009419:Nerve Tissue Proteins; D012710:Serum Albumin, Bovine