Quantitation of hepatitis B virus DNA in serum by ammonium sulfate precipitation and molecular hybridization
作者:
Long T. Wen,
Michael Henneberger,
Nicole Nguyen,
Richard A. McPherson,
期刊:
Journal of Clinical Laboratory Analysis
(WILEY Available online 1994)
卷期:
Volume 8,
issue 1
页码: 44-50
ISSN:0887-8013
年代: 1994
DOI:10.1002/jcla.1860080109
出版商: Wiley Subscription Services, Inc., A Wiley Company
关键词: hepatitis B virus DNA quantitation
数据来源: WILEY
摘要:
AbstractWe developed a method to quantitate hepatitis B virus (HBV) DNA in serum by ammonium sulfate fractionation and DNA hybridization. Serum samples were precipitated with 45% saturated ammonium sulfate, resuspended in buffer, and spotted on a nylon membrane. Following denaturation in alkali, HBV DNA sequences on the membrane were detected by hybridization with a32P‐labeled DNA probe of the entire HBV genome. Bound radioactivity was measured with liquid scintillation counting. Ammonium sulfate fractionation of positive samples increased assay sensitivity by 10–30‐fold compared to no treatment. Sensitivity for detection of cloned HBV DNA added to negative serum was 0.2 pg. Recovery of cloned HBV DNA added to negative serum before fractionation was equivalent to direct spotting of DNA onto the membrane in the absence of serum. This method enhanced HBV DNA recovery from serum into small volumes, thereby expanding the potential analytic range of spot hybridization assays. © 1994 Wiley‐L
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