Wirth S, Schaefer E, Winterpacht A, Zabel B
Kinderklinik, Johannes-Gutenberg-Universität, Mainz.
Monatsschr Kinderheilkd. 1992 Oct;140(10):759-62.
Detection of hepatitis B virus DNA is a reliable evidence of the presence of the viral agent and its replication. Conventional hybridization techniques are limited to detect about 30,000 virions. With the polymerase chain reaction it became possible to extend the sensitivity by amplification of viral sequences. In our study we intended to test whether viral sequences could be found in liver tissue specimens negative for hepatitis B virus DNA by conventional hybridization techniques.
Hepatitis B virus DNA was detected by PCR in liver tissue of 37 children with chronic hepatitis B, negative for hepatitis B virus DNA by Southern blot hybridization. PCR was performed in a thermal cycler using Taq-polymerase and oligonucleotide primers within the hepatitis B core region. Hepatitis B virus DNA was visualized by ethidium bromide staining and subsequent Southern blot hybridization.
20 patients were HBeAg- and 17 anti-HBe-seropositive. Viral sequences were present in each of the 20 HBeAg positive HBsAg carriers and in 10 patients with anti-HBe. No hepatitis B virus DNA could be found in 7 children, all of them positive for anti-HBe.
Our results confirm polymerase chain reaction to be a more sensitive method to detect hepatitis B virus DNA in the liver compared with conventional hybridization techniques. Every HBeAg positive carrier as well as the majority of anti-HBe positive patients present viral DNA in their liver. Polymerase chain reaction will be suitable to monitor viral replication in spontaneous course and treated patients.
检测乙型肝炎病毒DNA是病毒病原体存在及其复制的可靠证据。传统杂交技术检测病毒粒子的能力有限，约为30,000个。聚合酶链反应使通过扩增病毒序列来提高检测灵敏度成为可能。在我们的研究中，我们旨在测试在传统杂交技术检测乙型肝炎病毒DNA呈阴性的肝组织标本中是否能找到病毒序列。
采用聚合酶链反应（PCR）检测37例慢性乙型肝炎儿童肝组织中的乙型肝炎病毒DNA，这些儿童经Southern印迹杂交检测乙型肝炎病毒DNA呈阴性。在热循环仪中使用Taq聚合酶和乙型肝炎核心区域内的寡核苷酸引物进行PCR。通过溴化乙锭染色及随后的Southern印迹杂交对乙型肝炎病毒DNA进行可视化分析。
20例患者HBeAg阳性，17例抗-HBe血清学阳性。20例HBeAg阳性的HBsAg携带者及1例抗-HBe阳性患者的肝组织中均存在病毒序列。7例儿童未检测到乙型肝炎病毒DNA，他们均为抗-HBe阳性。
我们的结果证实，与传统杂交技术相比，聚合酶链反应是一种检测肝组织中乙型肝炎病毒DNA更灵敏的方法。每个HBeAg阳性携带者以及大多数抗-HBe阳性患者的肝脏中均存在病毒DNA。聚合酶链反应将适用于监测自然病程及治疗患者中的病毒复制情况。
