Cubie H A, Molyneaux P J, Shearman M J, Gryzbowski J, Brown T
Regional Virus Laboratory, City Hospital, Edinburgh.
Mol Cell Probes. 1995 Feb;9(1):59-65. doi: 10.1016/s0890-8508(95)91037-9.
Pools of 10 synthetic oligonucleotides with sequences derived from the genome of parvovirus B19 and of 30 bases in length were made and labelled at the time of synthesis with digoxigenin (DIG) or dinitrophenyl (DNP) at the 5' end. They were used in a dot-blot hybridisation assay to detect parvovirus B19 DNA in sera submitted for routine virological diagnosis where parvovirus infection was suspected. Detection down to 10-100 fg DNA (equivalent to 10(3)-10(4) copies of parvovirus B19 genome) was obtained with both probe cocktails and colorimetric or chemiluminescent detection systems. Of 141 clinical samples examined from 126 patients presenting with rash and/or joint pains, 107 were clearly negative with both probes, 20 were clearly positive and the remaining 14 samples gave discrepant results. Of these 34 samples, 33 contained parvovirus B19 specific IgM. The parvovirus oligonucleotide probe cocktail produced and labelled with either DIG or DNP provided a useful diagnostic reagent for the detection of specific DNA in clinical specimens using a simple and sensitive dot-blot assay.
制备了10组合成寡核苷酸池，其序列源自细小病毒B19基因组，长度为30个碱基，并在合成时于5'端用洋地黄毒苷（DIG）或二硝基苯基（DNP）进行标记。它们用于斑点印迹杂交试验，以检测提交进行常规病毒学诊断且怀疑有细小病毒感染的血清中的细小病毒B19 DNA。使用两种探针混合物以及比色或化学发光检测系统，均可检测到低至10 - 100 fg的DNA（相当于10³ - 10⁴份细小病毒B19基因组拷贝）。在对126例出现皮疹和/或关节疼痛的患者的141份临床样本进行检测时，107份样本用两种探针检测均明显为阴性，20份样本明显为阳性，其余14份样本结果不一致。在这34份样本中，33份含有细小病毒B19特异性IgM。用DIG或DNP制备并标记的细小病毒寡核苷酸探针混合物，为使用简单且灵敏的斑点印迹试验检测临床标本中的特异性DNA提供了一种有用的诊断试剂。
