Development of a measles specific IgM ELISA for use with serum and oral fluid samples using recombinant measles nucleoprotein produced in Saccharomyces cerevisiae.
In order to develop sensitive assays for detecting measles antibodies in oral fluid specimens, we have produced recombinant measles virus nucleoprotein (rMVN) in a yeast expression system and prepared monoclonal antibodies to the protein. Measles nucleoprotein gene from the Schwarz vaccine strain was cloned into a yeast expression vector, pFX7 under the control of the hybrid GAL10-PYK1 promoter. High levels of rMVN (20 mg/litre of yeast culture) were generated. Electron microscopy showed that the purified rMVN assembled into typical herring-bone structures. Monoclonal antibodies produced to the rMVN also reacted with native measles virus N in immunofluorescence tests. The purified rMVN and a monoclonal antibody to the rMVN conjugated to horseradish peroxidase were used to develop a measles specific IgM capture EIA (MACEIA) in both serum and oral fluid specimens. Evaluations of the MACEIA were performed by testing a) serum samples (n=80) and b) paired oral fluid/serum samples from measles cases (n=50, representing 16 cases) and oral fluids from controls with non-measles rash (n=59, representing 48 cases). The samples were also tested for measles IgM, using a reference radioimmunoassay (MACRIA). The sensitivity and specificity of the MACEIA compared with MACRIA for a) the serum samples were 100 and 96.6% respectively and b) for paired serum/oral fluids samples 100 and 100%, respectively.
Samuel
Dhanraj
D
Enteric, Respiratory and Neurological Virus Laboratory, Central Public Health Laboratory, 61 Colindale Avenue, London NW9 5HT, UK. dsamuel@phls.nhs.uk
Sasnauskas
Kestutis
K
Jin
Li
L
Gedvilaite
Alma
A
Slibinskas
Rimas
R
Beard
Stuart
S
Zvirbliene
Aurelija
A
Oliveira
Solange Artimos
SA
Staniulis
Juozas
J
Cohen
Bernard
B
Brown
David
D
D000818:Animals; D000914:Antibodies, Viral; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007075:Immunoglobulin M; D008457:Measles / Q000175:diagnosis; D008459:Measles virus / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009698:Nucleoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D012463:Saliva / Q000276:immunology; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral; D007075:Immunoglobulin M; D009698:Nucleoproteins; D011994:Recombinant Proteins