Dependence of the murine antibody response to an anti-CDR2 VH peptide on immunogen formulation.
A peptide corresponding to the second complementarity determining region of the heavy chain (CDR2 VH) from a murine anti-CD4 monoclonal antibody, designated L202, was synthesized by solid phase methodology in a number of different antigenic forms, for the purpose of comparing the effectiveness of different adjuvant-carrier systems in the induction of a murine antibody response against the immunizing peptide and parent antibody molecule. Two of the synthetic constructs contained the palmitoyl and N-palmitoyl-cysteinyl-S-(2,3-palmitoyloxy)-propanediol (PAM3Cys) moieties, respectively, attached to the peptide amino terminus with the immunogen comprising liposomal formulations of each. A third immunogen consisted of the CDR2 VH peptide admixed with the PAM3Cys non-covalently and incorporated into liposomes (PAM3Cys + CDR2 VH). A fourth composition comprised the CDR2 VH peptide conjugated to KLH via the sulfhydryl of an added N terminal cysteine (KLH-CDR2 VH) and injected with Complete Freund's adjuvant (CFA). A fifth immunogen consisted of the CDR2 VH peptide synthesized on an octameric, branched polylysine core as a multiple antigenic peptide (MAP-CDR2 VH) injected in the presence of Freund's adjuvant. Groups of five mice were injected intramuscularly with each of these immunogens and bled at two week intervals. The highest anti-peptide gamma-immunoglobulin (IgG) responses (against uncoupled peptide by ELISA) after 56 days were obtained with mice receiving the PAM3Cys-CDR2 VH peptide. However, when screened against the CDR2 V(H) peptide present as the MAP derivative by ELISA, IgG raised against the cognate MAP-CDR2 peptide was much more reactive than IgG raised against the liposomal PAM3Cys-CDR2 VH immunogen. In either case, IgG raised against the KLH-CDR2VH conjugate was poorly reactive. These differences in reactivity to the two forms of the CDR2 VH peptide by ELISA did not correspond to major differences in reactivities to the intact L202 Ab by ELISA. Although the IgG against the MAP immunogen was slightly more reactive than the other antisera against the l202 Ab, all titers were less than 1:100. These data illustrate some limitations of using anti-peptide responses as indicators of potential reactivity against the native protein, but suggest that alternate formulations including lipoidal peptides are more effective than corresponding KLH-peptide conjugates in eliciting Ab responses against poorly immunogenic epitopes.
Kanda
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Department of Virology and Immunology, Southwest Foundation for Biomedical Research, San Antonio, TX 78228-0147, USA.
Fritz
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DA
Gage
D A
DA
Shuler
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KR
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000737:chemistry; D015704:CD4 Antigens / Q000737:chemistry / Q000276:immunology*; D016056:Immunodominant Epitopes / Q000737:chemistry* / Q000276:immunology*; D007074:Immunoglobulin G / Q000096:biosynthesis*; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000737:chemistry*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000737:chemistry*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000276:immunology*; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D015704:CD4 Antigens; D016056:Immunodominant Epitopes; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D010455:Peptides