Derivation of monoclonal antibodies to human somatomedin C/insulin-like growth factor I.
Somatomedin C, also called insulin-like growth factor I (Sm-C/IGF-I), is a highly conserved polypeptide required for the proliferation of many cell types. Since several attempts in our laboratory to recover monoclonal antibody-secreting hybrids to this peptide by the direct fusion of hyperimmunized splenocytes with myeloma cells had been unsuccessful, we modified our approach by coculturing hyperimmunized BALB/c splenocytes and a small amount of the antigen for 5 days prior to fusion with the P3X63Ag.8.653 myeloma cell line. Of 88 microcultures at risk, specific antibody was detected in 24. Two clones were expanded in ascites fluid and characterized as to isotype, affinity, and specificity. Both were IgG1,kappa and bound human Sm-C/IGF-I with affinity constants of 1.09 and 1.01 X 10(10) liter/mol, respectively. Both clones were quite specific for Sm-C/IGF-I with inconsequential binding to insulin-like growth factor II, multiplication-stimulating activity, any of the chymotryptic fragments of Sm-C/IGF-I, insulin preparations, hGH, hTSH, mEGF, or mouse albumin. In vitro boosting after primary in vivo immunization appears to provide monoclones of an IgG isotype in contrast to primary in vitro immunization, which reportedly favors an IgM isotype. The antibodies produced in this study have proved to be extraordinarily useful in defining the physiologic role of Sm-I/IGF-I with immunoneutralization techniques and in the purification of human Sm-C/IGF-I by affinity chromatography.
Gillespie
G Y
GY
Van Wyk
J J
JJ
Underwood
L E
LE
Svoboda
M E
ME
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000302:isolation & purification; D007136:Immunoglobulins; D007334:Insulin-Like Growth Factor I / Q000032:analysis / Q000276:immunology*; D007457:Iodine Radioisotopes; D013002:Somatomedins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007136:Immunoglobulins; D007457:Iodine Radioisotopes; D013002:Somatomedins; D007334:Insulin-Like Growth Factor I