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本研究应用国产生物素标记HBVDNA探针,于福尔马林固定,石蜡包埋肝组织切片上,建立了碱性磷酸酶过氧化物酶显色的原位杂交技术,在此过程中发现,蛋白酶的消化是杂交能否成功的关键因素,酸、三乙醇酐、洋地黄的预处理,能促进探针参入,降低非特异背景,减低蛋白酶作用浓度或作用时间,因而提高实验稳定性、特异性。并用这一技术检测了34例肝活检和4例尸检肝组织,结果:尸肝组织均检出了HBV—DNA,其中2例核型见于肝硬化,肝癌旁组织,而2例浆型HBV—DNA见于慢活肝,亚重肝。34例肝活检组织中,只检出12例浆型HBV—DNA,其中CAH—CPH无症状携带者中检出率分别为42.9％,38.8％,33.3％,均明显高于肝硬化组织(16.7％)(P<0.05)。由此可见浆型HBV—DNA分布同病变轻重程度的关系可能并不密切,不同分布型的HBV—DNA可能只反映HBV的复制状态。
In this study, domestic biotin-labeled HBV DNA probes were used to formalin-fixed, paraffin-embedded liver tissue sections, and an in situ hybridization technique for alkaline phosphatase peroxidase staining was established. Protease digestion is a key factor in the success of hybridization. Pretreatment with acid, triethanol anhydride, and digitalis can promote the incorporation of probes, reduce non-specific background, and reduce the concentration or time of protease action, thus improving the stability and specificity of the assay. Sex. And this technique was used to detect 34 cases of liver biopsy and 4 cases of autopsy liver tissue. Results: HBV-DNA was detected in cadaveric liver tissues. Two cases of karyotype were found in cirrhosis and adjacent tissues of hepatocellular carcinoma, and 2 cases of plasma HBV- DNA is found in slow-lived liver, sub-hepatic liver. Of the 34 liver biopsies, only 12 cases of plasma HBV-DNA were detected, and the detection rates of asymptomatic CAH-CPH carriers were 42.9%, 38.8%, and 33.3%, respectively, which were significantly higher than those of cirrhosis (16.7). %) (P<0.05). It can be seen that the relationship between plasma HBV-DNA distribution and the degree of severity of the lesion may not be closely related, different distribution of HBV-DNA may only reflect the replication status of HBV.