Monoclonal antibody that neutralizes pertussis toxin activities.

Pertussis or whooping cough is a disease with high mortality among infants and small children. The disease is caused by infection of the respiratory tract by a gram negative bacterium, Bordetella pertussis. The superficial colonized bacteria produce a myriad of toxins which enter the circulation causing various pathophysiologicalal changes in the host. Although antimicrobial therapy reduces the number of the coughed out bacteria and also the infectious time of the infected host, but it is not effective in amelioration of the clinical manifestations as the pertussis morbidity is due principally to the pertussis toxin (PT). Antibody based-therapy is frequently practiced in conjunction with other supportive measure to resuscitate the patient. Nevertheless, human derived antiserum against PT is of the limited supply and the ethical concern. Thus in this study a hybridoma clone, i.e. clone PT6-2G6, secreting monoclonal antibody (MAb) specific to the S1 subunit, the active enzyme of the PT that intracellularly ADP-ribosylates the host Gi-protein, was produced. The MAbPT6-2G6 inhibited the in vitro hemagglutination of chicken erythrocytes which is the activity of the B oligomer of PT; thus we hypothesize that the MAb bound to its epitope on the S1 subunit and stereologically hinders the binding sites of the B subunits. The MAb also inhibited ex vivo Chinese hamster ovarian cell clustering and neutralized the in vivo leucocytosis- promotion in mice which are usually mediated by intracellular S1 subunit. The large molecular nature of the intact MAb and its molecular hydrophilicity led us to speculate that the observed PT neutralizing activities of the MAb were due to interfering with the cellular entry of the S1 rather than the intracellular enzyme neutralizing activity per se. While further experiments are needed to pinpoint the MAb neutralizing activity and to identify the amino acid sequence and location of the MAbPT6-2G6 epitope, our findings indicate that this murine MAb, in its humanized-version, should have high therapeutic potential for pertussis.

Rendimento, tempo de destoxificacao e eficiencia do processo de producao da Vacina Pertussis com os meios de cultura de Cohen & Wheeler e Stainer & Scholte / Yield, time of detoxification and efficiency of the production of Pertussis Vaccine in Cohen & Wheeler and Stainer & Scholte culture media

Cultivos de 24 horas de Bordetella pertussis em fermentadores Biolafitte 50L, obtidos em meios de cultura de STAINER & SCHOLTE (SS) e COHEN & WHEELER (CW), foram inativados por formalina 0,1% a 35 C por 48 horas e a 25 C por 24 horas, ou pelo calor a 56 C por 30 minutos. As suspencoes foram, entao, concentradas por centrifugacao. A utilizacao do meio CW para o cultivo de B.pertussis seguido de inativacao por formalina 0,1%, demonstrou ser o metodo de producao da Vacina Pertussis mais adequado, tendo em vista o rendimento global, o custo de producao e o tempo necessario a destoxificacao. O meio de CW apresentou uma economia da ordem de 41,28% em relacao ao meio SS, quando avaliados os custos de producao. Apotencia das vacinas foram superiores a 4UI/dose e demonstraram uma correlacao significante entre atividade imunogenica e Fator Promotor da Leucocitose (LPF).