Urdea M S, Running J A, Horn T, Clyne J, Ku L L, Warner B D
Chiron Corporation, Emeryville, CA 94608.
Gene. 1987;61(3):253-64. doi: 10.1016/0378-1119(87)90189-2.
The detection of a little as 0.2 pg (60,000 molecules) of hepatitis B viral (HBV) DNA in human serum samples in 4 h has been demonstrated using a solution-hybridization and bead-capture method. An amplification method based on chemically crosslinked oligodeoxyribonucleotides was coupled with a horseradish peroxidase-labeling scheme for the ultimate detection of the analyte. Two sets of HBV complementary synthetic oligodeoxyribonucleotide probes containing one of two types of single-stranded (ss) overhangs were employed. These ss overhangs were used to capture the probe-analyte complex onto a bead and subsequently to label it. Detection was achieved with either a chemiluminescent or colorimetric output substrate for the enzyme. Only in the presence of the virus was label specifically bound to the support. The assay was relatively unaffected by either sample composition or by the presence of heterologous nucleic acids.
采用溶液杂交和磁珠捕获法，已证实在4小时内可检测出人血清样本中低至0.2皮克（60,000个分子）的乙型肝炎病毒（HBV）DNA。基于化学交联寡脱氧核糖核苷酸的扩增方法与辣根过氧化物酶标记方案相结合，用于最终检测分析物。使用了两组含有两种单链（ss）突出端之一 的HBV互补合成寡脱氧核糖核苷酸探针。这些ss突出端用于将探针 - 分析物复合物捕获到磁珠上，随后对其进行标记。使用酶的化学发光或比色输出底物进行检测。只有在病毒存在的情况下，标记物才会特异性结合到载体上。该检测方法相对不受样品组成或异源核酸存在的影响。
