A quantitative immuno-PCR assay for the detection of mumps-specific IgG.
Sensitive assays are required for seroprevalence studies of measles, mumps and rubella (MMR)-vaccinated populations where many may have low levels of antibodies. This protocol describes a quantitative immuno-PCR assay to detect mumps-specific IgG antibodies. The purpose of the protocol is to determine the immune status of individuals to mumps. Mumps-specific IgG from a dilution of patients serum is bound by recombinant mumps nucleoprotein coated on the surface of microtitre plate wells. Bound antibody is detected by PCR using a conjugate of anti-human IgG covalently coupled to an oligonucleotide. The oligonucleotide is detected by the addition of target DNA, designed to hybridise to the oligonucleotide and serve as a template for real-time PCR using the LightCycler. The quantity of target DNA detected by the PCR depends upon the level of specific antibody in the test sample.
McKie
Anne
A
Genomics, Proteomics and Bioinformatics Services, Central Public Health Laboratory, 61 Colindale Avenue, Colindale, London NW9 5HT, UK.
Samuel
Dhanraj
D
Cohen
Bernard
B
Saunders
Nicholas A
NA
D000914:Antibodies, Viral / Q000097:blood* / Q000235:genetics / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood* / Q000235:genetics / Q000276:immunology; D009107:Mumps / Q000097:blood / Q000821:virology*; D009109:Mumps virus / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000276:immunology*; D009698:Nucleoproteins / Q000235:genetics / Q000276:immunology*; D016133:Polymerase Chain Reaction / Q000379:methods*
D000914:Antibodies, Viral; D007074:Immunoglobulin G; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins