G-protein based ELISA as a potency test for rabies vaccines

Abstract

The NIH test is currently used to assess the potency of rabies vaccine, a key criterion for vaccine release. This test is based on mice immunization followed by intracerebral viral challenge. As part of global efforts to reduce animal experimentation and in the framework of the development of Sanofi Pasteur next generation, highly-purified vaccine, produced without any material of human or animal origin, we developed an ELISA as an alternative to the NIH test. This ELISA is based on monoclonal antibodies recognizing specifically the native form of the viral G-protein, the major antigen that induces neutralizing antibody response to rabies virus. We show here that our ELISA is able to distinguish between potent and different types of sub-potent vaccine lots. Satisfactory agreement was observed between the ELISA and the NIH test in the determination of the vaccine titer and their capacity to discern conform from non-conform batches. Our ELISA meets the criteria for a stability-indicating assay and has been successfully used to develop the new generation of rabies vaccine candidates. After an EPAA international pre-collaborative study, this ELISA was selected as the assay of choice for the EDQM collaborative study aimed at replacing the rabies vaccine NIH in vivo potency test.

Development and Evaluation of an Immuno-Capture Enzyme-Linked Immunosorbent Assay to Quantify the Mycoplasma capricolum subsp. capripneumoniae (Mccp) Protein in Contagious Caprine Pleuropneumonia (CCPP) Vaccine

“…Seven repeated quantification tests were performed, and a 2 value between 0.95 and 0.98 was obtained, indicating a good relationship between the amount of protein tested in the ICE and the OD values obtained. is is similar to the correlation ( 2 ) reported [32] in G-protein based ELISA as potency test for rabies vaccine. amount of protein required (0.150 mg/ml) [34].…”

Rabies Vaccine Characterization by Nanoparticle Tracking Analysis

“…All samples were assessed with or without anti-G D1-25 to the rabies glycoprotein conformational epitope, as previously described 25 . In brief, for the anti-G D1-25 labelled particles, RABV samples were incubated for 1 hour at 37 °C with anti-G D1-25 monoclonal antibody directly conjugated to Alexa Fluor 488 by BIOTEM (Apprieu, France), at 1:200 dilution in phosphate buffered saline.…”

“…Plates were then washed and buffer containing revelation substrate added to each well. The OD 492 nm was determined using an ELISA reader (Molecular Devices) as previously described 25 . This ELISA was proposed to the European Directorate for the Quality of Medicines and Healthcare (EDQM) and accepted for an international collaborative study to replace the NIH activity tests for rabies vaccines 32 .…”

Overcoming scientific barriers in the transition fromin vivoto non-animal batch testing of human and veterinary vaccines

“…To solve these problems, vaccine batches could be altered intentionally, for example by exposure to stresses that may decrease the stability of a batch during the vaccine production process (e.g. decreased or increased pH, osmolality, temperature), by creating vaccines with reduced antigen content or a different composition, by deviating from the standard inactivation method, or by changing the degree of adsorption to adjuvants [11,34,41,57,124,126,127]. Importantly, altered batches should be representative for non-compliant batches that may realistically be produced as a result of disturbances in vaccine production processes.…”