Quantitative RT-PCR ELISA to determine the amount and ratio of positive- and negative strand viral RNA synthesis and the effect of guanidine in poliovirus infected cells.
Quantitative reverse transcription-polymerase chain reaction enzyme linked immunosorbent assay (RT-PCR ELISA) is the method of choice to study positive- and negative strand viral RNA synthesis during poliovirus replication. In comparison with other methods used for this purpose, it fulfils all necessary requirements to accurately determine RNA of different polarity. It combines high specificity, high sensitivity, safety, speed, and the ability to perform quantitative analysis. The enterovirus specific RT-PCR ELISA method described in this work, was used to determine quantitatively the amount of de novo poliovirus positive- and negative strand RNA synthesis at different time-points in the viral replication cycle, both in presence and absence of the viral RNA synthesis inhibitor guanidine hydrochloride.
Verheyden
B
B
Department of Microbiology and Hygiene, Vrije Universiteit Brussel, Laarbeeklaan 103, B-1090 Brussels, Belgium.
Lauwers
S
S
Rombaut
B
B
D001710:Biotin / Q000737:chemistry; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D019791:Guanidine / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D011051:Poliomyelitis / Q000378:metabolism* / Q000821:virology; D017955:Poliovirus; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019809:Streptavidin / Q000737:chemistry
D017931:DNA Primers; D012367:RNA, Viral; D001710:Biotin; D019809:Streptavidin; D019791:Guanidine