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Induction of polyclonal antibodies to the S1 subunit of pertussis toxin by synthetic peptides coupled to PPD: Effect of conjugation method, adjuvant, priming and animal species
Two peptides, designated L and K, covering a sequence near the NH‐terminal end of the S1 subunit of pertussis toxin (PT) were conjugated to the PPD (purified protein derivative) of M. tuberculosis by either glutaraldehyde (GLUT) or succinimidyl 4‐(N‐maleimidomethyl) cyclohexane‐1‐carboxylate (SMCC) and N‐succinimidyl 3‐(2‐pyridyldithio) propionate (SPDP) and injected into groups of mice and guinea pigs. Initially, the effect of priming the animals with BCG vaccine and the use of aluminium hydroxide as adjuvant for the anti‐peptide antibody response was studied. The group of BCG‐primed mice immunized with adsorbed peptide conjugates showed the highest anti‐peptide conjugate antibody response. Based on this finding, groups of BCG‐primed mice were immunized four times with either adsorbed peptide L‐GLUT, peptide L‐SMCC/SPDP or peptide K‐SMCC/SPDP conjugates and the fine peptide specificity as well as the PT and SI cross‐reactivity was investigated in ELISA. Mice immunized with peptide L‐GLUT showed a significant antibody response to the homologous conjugate, only, whereas the group injected with the peptide L‐SMCC/SPDP conjugate gave a significant response to both peptide K and L conjugated by the SMCC‐SPDP method. Likewise, mice immunized with the peptide K‐SMCC/SPDP conjugate reacted with the homologous and peptide L‐SMCC/SPDP conjugate, although only the response to the former conjugate was significantly greater than the response to PPD. All groups showed a strong anti‐PPD response. The anti‐PT/S1 cross‐reactivity of the antisera varied considerably within each group but was found to be highest in the peptide L‐GLUT‐immunized animals. The results of the present study not only stress the importance of BCG priming and use of aluminium hydroxide adjuvants for the immunogenicity of the peptides in question but also point to the specificity of the conjugation methods employed as low cross‐reactivity between the anti‐peptide L‐GLUT and L‐SMCC/SPDP antisera was noted. Moreover, it appeared that the choice of conjugation method may have an effect on the ability of the peptide conjugates to induce an antibody response cross‐reacting with the native protein.