Influence of fluorescent antibody probe specificity on flow cytometric analysis of antibody-producing cells.
In the flow cytometric analysis of the stability of antibody-producing cells, fluorescent antibody probes specific for immunoglobulin heavy chains have been widely utilized to quantify intracellular antibodies. To investigate the effect of the specificity of antibody probes on flow cytometric analysis, non-producing subclones of the 6-31 transfectoma were stained with fluorescein isothiocyanate (FITC)-conjugated anti-human IgGs specific for heavy chain and light chain, respectively. The use of heavy chain-specific probe identified heavy chain-only producers as producers, whereas the use of light chain-specific probe identified light chain-only producers as producers. Thus, both heavy chain-specific and light chain-specific antibody probes should be used for the accurate evaluation of heterogeneous non-producing population. Furthermore, the results of the flow cytometric analysis were confirmed by immunoblotting, suggesting that flow cytometry is a useful technique for the rapid evaluation of the stability of transfectomas producing chimeric antibody.
Bae
S W
SW
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Taejon, South Korea.
Lee
G M
GM
D000818:Animals; D000918:Antibody Specificity; D000921:Antibody-Producing Cells / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D015342:DNA Probes; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D006825:Hybridomas / Q000378:metabolism; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics; D007145:Immunoglobulin kappa-Chains / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014162:Transfection
D015342:DNA Probes; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains; D011993:Recombinant Fusion Proteins