RESUMEN
This study addresses observations made in view of testing in practice the guideline in the European Pharmacopoeia (EP) on omitting the rat potency test for release of polio containing vaccines. In general, use of the guideline is valid and the D-antigen ELISA can indeed be used as an in vitro alternative for the in vivo test. However, the set-up of the ELISA is crucial and should include detection of antigenic site 1 in polio serotype 3 as destruction of that site by trypsin results in a reduced rat potency. Antigenic site 1 in polio serotype 2 may also be modified by trypsin, but the cleavage of viral protein 1 (VP1) did not affect the rat potency. Therefore, any antigenic site, except site 1, can be used for detection of polio serotype 2. It is advised to include testing of the effect of trypsin treatment in the EP-guideline. This allows polio vaccine manufacturers to check whether their in-house ELISA needs improvement.
Asunto(s)
Bioensayo , Ensayo de Inmunoadsorción Enzimática , Vacuna Antipolio de Virus Inactivados/inmunología , Poliovirus/efectos de los fármacos , Tripsina/farmacología , Animales , Anticuerpos Neutralizantes/biosíntesis , Anticuerpos Antivirales/biosíntesis , Técnicas Biosensibles , Proteínas de la Cápside/efectos de los fármacos , Chlorocebus aethiops , Relación Dosis-Respuesta Inmunológica , Europa (Continente) , Calor , Poliovirus/inmunología , Poliovirus/aislamiento & purificación , Poliovirus/patogenicidad , Guías de Práctica Clínica como Asunto , Ratas , Células Vero , Virulencia/efectos de los fármacosRESUMEN
A fast ELISA was developed and qualified for analysis of polio D-antigen. The original 20 h-protocol was optimized by minimizing the total incubation time to 1 h, and by replacing the signal reagent 3,3',5,5'-tetramethylbenzidine by a chemiluminogenic signal reagent with a theoretical low intrinsic background and high dynamic range.
