Application of flow cytometric measurement of surface IgG in kinetic analysis of monoclonal antibody synthesis and secretion by murine hybridoma cells.
The kinetics of antibody synthesis and secretion from a murine hybridoma cell line were studied using measurements of total cell-associated IgG, surface IgG, and IgG secreted into the medium. Kinetic analysis of IgG secretion demonstrates approximately constant secretion rate per viable cell over the entire batch cultivation. A correlation was observed (r2 = 0.74) between mean surface immunofluorescence and the total cell-associated IgG determined by ELISA of detergent-extracted cell lysates. No correlation was found between specific secretion rate and mean surface IgG level estimated by immunofluorescence flow cytometry measurements. Material balances on cellular IgG demonstrated that about 7% of the antibody which was synthesized during exponential batch growth was not released to the growth medium. Distributions of single-cell surface antibody content showed two subpopulations, one with very low surface IgG. The fraction of the population with low surface IgG increased throughout a batch cultivation.
Department of Chemical Engineering, California Institute of Technology, Pasadena 91125.
E
E
Meilhoc
K D
KD
Wittrup
J E
JE
Bailey
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D002460:Cell Line; D005434:Flow Cytometry; D006825:Hybridomas / Q000378:metabolism*; D007074:Immunoglobulin G / Q000032:analysis*; D051379:Mice; D011947:Receptors, Antigen, B-Cell / Q000032:analysis*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D011947:Receptors, Antigen, B-Cell