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目的检测BRAF基因在黑素瘤组织(MM)中的表达情况,并探讨运用实时荧光定量检测石蜡包埋组织中BRAF mRNA表达的方法。方法收集5例MM标本分别经福尔马林固定石蜡包埋和直接冻存提取RNA,以5例正常皮肤组织为正常对照组,采用实时荧光定量PCR分析BARF基因的mRNA表达情况,并与免疫组织化学的结果进行对比。结果冰冻、石蜡包埋的MM中BRAF mRNA平均表达水平(0.267 0±0.101 1)显著高于正常皮肤组织中BRAF平均表达水平(0.174 1±0.016 1),差异具有统计学意义(P<0.05)。冰冻与石蜡包埋组织BRAF的相对表达量RQ值结果一致,免疫组化的结果显示BRAF在MM中表达明显高于正常对照组。结论可以利用实时荧光定量对石蜡包埋组织进行RNA相关表达研究,与新鲜组织的结果一致性好,并与免疫组化结果一致。
Objective To detect the expression of BRAF gene in melanoma (MM) and to explore the method of real-time fluorescence quantitative detection of BRAF mRNA expression in paraffin-embedded tissue. Methods Five specimens of MM were collected and formalin-fixed paraffin-embedded and cryopreserved respectively for RNA extraction. Five normal skin tissues were used as normal control group. MRNA expression of BARF gene was analyzed by real-time fluorescence quantitative PCR. Histochemistry results were compared. Results The average expression level of BRAF mRNA in frozen and paraffin-embedded MM was significantly higher than that in normal skin (0.267 0 ± 0.101 1, 0.174 1 ± 0.016 1, P <0.05) . The results of RQ showed that the expression of BRAF in frozen and paraffin-embedded tissues was consistent with the results of immunohistochemistry, and the expression of BRAF in MM was significantly higher than that in normal controls. Conclusion Real-time fluorescence quantitative analysis of RNA-dependent expression of paraffin-embedded tissues is consistent with the results of fresh tissues and is consistent with the results of immunohistochemistry.