Quantitative nested PCR for human cytomegalovirus DNA: analysis using NucleoScan 2001.
A quantitative nested DNA PCR for human cytomegalovirus (CMV) was developed using PCR mimic DNA as an internal standard. A combination of ultra-thin gel and enhanced detection of ethidium bromide-stained PCR bands allowed us to achieve a 25-fold increase in sensitivity over the conventional gel electrophoreses, down to 0.1 ng of DNA per band. When the technique was applied to measuring CMV load in CMV positive leucocyte DNA preparations its sensitivity spanned the range from single to hundreds of CMV DNA copies in 0.1 microg of total white blood cell DNA.
Boriskin
Y S
YS
Department of Medical Microbiology, St George's Hospital Medical School, London, UK.
D003587:Cytomegalovirus / Q000235:genetics* / Q000302:isolation & purification; D003586:Cytomegalovirus Infections / Q000175:diagnosis*; D017931:DNA Primers / Q000737:chemistry; D004279:DNA, Viral / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D004996:Ethidium / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D007091:Image Processing, Computer-Assisted; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D017931:DNA Primers; D004279:DNA, Viral; D005456:Fluorescent Dyes; D004996:Ethidium