Antigenic markers of V domain of mouse MOPC 21 IgG₁ L chain. The conformational basis of V_L domain markers and content of MOPC 21 V_L‐like immunoglobulins in sera of inbred mouse strains

Abstract

Antisera were raised in rabbits against L chains, isolated from mouse myeloma protein MOPC21 (gamma 1, chi V chi 15 group). Specific antibodies for the V and C domain of MOPC21 L chain were obtained by cross-immunoadsorption of the antisera. The pure anti-V and anti-C antibodies were fixed on diazocellulose and used as immunosorbents. The inhibitory capacity of L chin-monomers and dimers isolated from the L chain preparation was compared to that of intact IgG1 using binding inhibition of 125I-labeled IgG1 on the antibody-containing immunosorbents. It was established that changes of IgG1 quaternary structure influences the conformational state of the L chain V domain only. The inhibitory capacity of the V domain is 1000-fold lower in L monomers, if compared with native IgG1, and only 10-fold lower than in L dimers. The inhibiting capacity of the C domain, however, does not differ in L monomers and intact IgG1. Thus the conformational rigidity of the C domain co-exists with conformational flexibility of the V domain on the same polypeptide chain. We tried to estimate the content of MOPC21 V1-like normal IgG in mouse serum of 6 inbred strains using antibodies against the V1 domain. Data obtained by inhibition of radioimmunoadsorption, indicate that in C57BL/6 mice 0.08% of normal serum Ig carries a V1 region which is idiotypically related to the V1 of MOPC21. In serum Ig of BALB/c mice the percentage is 0.16.

Appearance of Neoantigen in Mouse IgG1 Upon Reduction of Interchain Disulfide Bridges: Assessment of Local and General Conformational Rearrangements Using Monoclonal Antibody and Small-angle X-ray Scattering

“…H and L chains of monoclonal and normal lg were isolated by reduction and alkylation of disulfide H-L linkages (10 mM dithiothreitol, 23 mM iodoacetamide for reduction and alkylation of monoclonal lg; 30 mM dithiothreitol, 70 mM iodoacetamide for polyclonal lg) followed by gel filtration on Sephadex G-100 equilibrated with 1M acetic acid (11). Downloaded by [Rutgers University] at 16:42 10 April 2015…”

“…Intact, reduced and reconstituted Ig were labelled using iodine monochloride method (14) with specific radioactivity between 30,000 and 2,000,000 cpmh1g. Conditions of iodination and methods of immunoadsorption were described in detail elsewhere (11). Radioactive pellets ofimmunoadsorbants were counted in gamma-spectrometer Minigamma 1275 (LKB, Finland).…”

“…Antisera were raised in rabbits against intact mouse Ig and isolated L chains. Pure monospecific rabbit Ab against V and C domains of murine Ig were isolated as described previously (11). Intact, reduced and reconstituted Ig were labelled using iodine monochloride method (14) with specific radioactivity between 30,000 and 2,000,000 cpmh1g.…”