A novel serological technique: polymerase chain reaction enhanced immunoassay. Application to enterovirus IgM diagnosis.
The polymerase chain reaction (PCR) method is a sensitive, specific and rapid technique for virus detection. The principles of a PCR enhanced immunoassay (PIA) are described. The method combines solid phase serological techniques with the PCR, providing a versatile and sensitive method for antibody detection. By linking the antigenicity of virus particles with their content of nucleic acid, the method provides new possibilities for virus serology: for example, antibody specificity can be coupled to viral sequence in patients with chronic infections caused by highly variable viruses such as HIV and HCV. An application of the PIA technique is described for the detection of anti-enterovirus IgM. IgM is captured to anti-human IgM-coated microwell plates. The anti-enterovirus IgM is allowed to bind crude enterovirus antigen. Bound virus is heat denatured and the released RNA is used as a template for reverse transcription PCR (RT-PCR) amplification. Amplicons are detected by hybridisation to an affinity labelled probe in a microwell colorimetric assay. In a pilot study, 18 serum specimens from patients with enterovirus infections were examined. Using a mixture of ten crude enterovirus antigens, the frequency of IgM positivity was 6/18 (33%). Titres between 1/500 and 1/100,000 were recorded. Predominantly type-specific antibodies were detected. The results were compared with a procapsid enterovirus radioimmunoassay (RIA). After further optimisation, the PIA has the potential to be a clinically useful assay for the detection of antiviral antibodies.
Aspholm
R
R
Department of Medical Sciences, Uppsala University Hospital, Sweden.
Zuo
S
S
Fohlman
J
J
Frisk
G
G
Friman
G
G
Blomberg
J
J
D000328:Adult; D000368:Aged; D002555:Cerebrospinal Fluid / Q000821:virology; D003384:Coxsackievirus Infections / Q000097:blood / Q000175:diagnosis; D004457:Echovirus Infections / Q000097:blood / Q000175:diagnosis; D004770:Enterovirus / Q000145:classification / Q000276:immunology* / Q000302:isolation & purification; D029822:Enterovirus B, Human / Q000145:classification / Q000276:immunology / Q000302:isolation & purification; D004769:Enterovirus Infections / Q000097:blood / Q000175:diagnosis*; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood; D008875:Middle Aged; D011863:Radioimmunoassay / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D007075:Immunoglobulin M