Qualitative PCR-ELISA protocol for the detection and typing of viral genomes.
PCR is an established technique providing rapid and highly productive amplification of specific DNA sequences. The demand for equally rapid, sensitive and objective methods to achieve detection of PCR products has led to the coupling of PCR with ELISA. PCR-ELISA involves direct incorporation of labeled nucleotides in amplicons during PCR-amplification, their hybridization to specific probes and hybrid capture-immunoassay in microtiter wells. PCR-ELISA is performed in 1 d and is very flexible, with the ability to process simultaneously up to 96 or 384 samples. This technique is potentially automatable and does not require expensive equipment, and thus can be fundamental in laboratories without access to a real-time PCR thermocycler. PCR-ELISA has mainly been used to detect infectious agents, including viruses, bacteria, protozoa and fungi. A PCR-ELISA protocol for the qualitative detection of papillomavirus genomes and simultaneous typing of different genotypes are detailed here as an example of the technique.
Musiani
Monica
M
Department of Clinical and Experimental Medicine, Division of Microbiology, University of Bologna, Via Massarenti 9, Bologna 40138, Italy. monica.musiani@unibo.it
Venturoli
Simona
S
Gallinella
Giorgio
G
Zerbini
Marialuisa
M
D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D016679:Genome, Viral; D027383:Papillomaviridae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers