Using a Sandwich ELISA to Determine the Immunogenic Glycoprotein Content in a Vaccine

Abstract

Source: Jallet, C. et al., In Vitro ELISA Test to Evaluate Rabies Vaccine Potency. J. Vis. Exp. (2020)

This video showcases an indirect enzyme-linked immunosorbent assay or ELISA sandwich immunocapture assay designed to assess immunogenic glycoprotein levels in vaccines. The assay plate, containing glycoprotein-specific antibodies, is incubated with serially diluted vaccine samples, including reference and test vaccines, followed by immunoassay to quantify glycoprotein content in the test vaccine.

Protocol

1. Security precautions NOTE: This method is applicable to both live rabies virus (RABV) and inactivated vaccine. Use good Laboratory Practice and Safety procedures. Wear adequate Personal Protection Equipment (PPE) including disposable coat, gloves, masks, glasses, etc. When the live virus is titrated, use a class II biological safety cabinet. Consider any material in contact with the samples (reagents, washing solu…

Materials

Class II Biological Safety Cabinet	ThermoFisher Scientific	10445753	If titrating live virus
Clear Flat-Bottom Immuno Nonsterile 96-Well Plates, 400 µL, MAXISORP	ThermoFisher Scientific	439454	Good for binding to the loaded antibody
Equip Labo Polypropylene Laboratory Fume Hood	ThermoFisher Scientific	12576606	For the preparation of sulfuric acid
Immunology Plate Strong Adsorption MAXISORP Flat Bottom Well F96	Dutscher	55303	Good for binding to the loaded antibody
Microplate Sealing Tape(100 sheets)	ThermoFisher Scientific	15036
Microplate  single mode reader Sunrise	TECAN
Microplate shaker-incubator	Dutscher	441504
Microplate washer Wellwash	ThermoFisher Scientific	5165000
Multichannel pipette (30-300 µL) 12 channels	ThermoFisher Scientific	4661180N
Single Channel pipettes (Kit 2 : Finnpipettes F2 0.2-2 μL micro, 2-20 μL, 20-200 μL & 100-1000 μL)	ThermoFisher Scientific	4700880