Cameron-Wilson Charlotte L, Muir Peter, Ballard Anna L, Corden Sally, Boxall Elizabeth H, Sablon Erwin, Stuyver Lieven
Department of Infectious Diseases, King's College London, St. Thomas' Campus, Lambeth Palace Road, London SE1 7EH, UK.
J Virol Methods. 2003 Dec;114(1):97-103. doi: 10.1016/j.jviromet.2003.09.014.
A prototype line probe assay (LiPA) for identifying hepatitis B virus (HBV) precore variants (INNO-LiPA HBV precore) was evaluated using a panel of 50 sera from 46 patients with HBV infection. The assay detected sequence variations detected commonly in the precore promoter region and in amino acid codons 28 and 29 of the precore gene. There was strong agreement between INNO-LiPA HBV precore results and those of a codon 28 point mutation assay (PMA), with identical results obtained in 40 of 43 sera (93%) typeable by both assays (kappa coefficient (kappa)=0.90). In addition, the precore codon 29 sequence identified by the INNO-LiPA HBV precore was confirmed by nucleotide sequencing in all seven samples analysed. However, the INNO-LiPA HBV precore identified precore promoter sequences much less efficiently. The prototype assay could identify codon 28/29 sequences from as little as 10 HBV genome equivalents in 10 microl serum, and in experiments using artificially prepared mixtures of variants could identify a minor component constituting 2.5% of the total viral DNA population. The INNO-LiPA HBV precore was also straightforward technically and rapid, and is therefore likely to be useful for epidemiological investigations into the prevalence, distribution and clinical significance of HBV precore variants.
使用来自46例乙肝病毒（HBV）感染患者的50份血清样本，对一种用于鉴定HBV前核心区变异的原型线性探针分析方法（LiPA）（INNO-LiPA HBV前核心区分析）进行了评估。该分析方法检测到了在前核心启动子区域以及前核心基因第28和29位氨基酸密码子中常见的序列变异。INNO-LiPA HBV前核心区分析结果与第28位密码子点突变分析（PMA）结果高度一致，在两种分析方法均可分型的43份血清样本中有40份（93%）结果相同（kappa系数（kappa）=0.90）。此外，在所有分析的7个样本中，通过核苷酸测序证实了INNO-LiPA HBV前核心区分析鉴定出的前核心第29位密码子序列。然而，INNO-LiPA HBV前核心区分析鉴定前核心启动子序列的效率要低得多。该原型分析方法能够从10微升血清中低至10个HBV基因组当量的样本中鉴定出第28/29位密码子序列，并且在使用人工制备的变异体混合物进行的实验中，能够鉴定出占病毒DNA总量2.5%的次要成分。INNO-LiPA HBV前核心区分析在技术上也很简单且快速，因此可能有助于对HBV前核心区变异的流行率、分布及临床意义进行流行病学调查。
