Automated detection of digoxigenin-labelled B19 parvovirus amplicons by a capture hybridization assay.
An automated method to identify B19 amplicons, directly labelled with digoxigenin during amplification reaction was developed. The labelled amplicons were hybridized with a biotinylated B19 oligo-probe and captured on commercially available test tubes coated with streptavidin. The hybridized amplicons labelled with digoxigenin were detected using anti-digoxigenin Fab fragments conjugated to peroxidase and the colourimetric reaction automatically evaluated as an immunoenzymatic assay. Fifty serum samples were tested by the assay and the results were in accordance with those obtained by Southern blot analysis of amplified products. Due to the high sensitivity, specificity and reproducibility shown, the assay seems to be a practical and reliable test for the diagnosis of B19 infection and can be easily adapted to identify any digoxigenin-labelled amplified product of viral genomes.
Zerbini
M
M
Institute of Microbiology, University of Bologna, Italy.
Gibellini
D
D
Musiani
M
M
Venturoli
S
S
Gallinella
G
G
Gentilomi
G
G
D001426:Bacterial Proteins / Q000737:chemistry; D001483:Base Sequence; D001710:Biotin / Q000737:chemistry; D015342:DNA Probes; D004279:DNA, Viral / Q000032:analysis* / Q000235:genetics; D004076:Digoxigenin; D016731:Erythema Infectiosum / Q000097:blood / Q000821:virology; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D008297:Male; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016732:Parvovirus B19, Human / Q000302:isolation & purification*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019809:Streptavidin
D001426:Bacterial Proteins; D015342:DNA Probes; D004279:DNA, Viral; D007140:Immunoglobulin Fab Fragments; D011933:Reagent Kits, Diagnostic; D001710:Biotin; D019809:Streptavidin; D004076:Digoxigenin