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[目的]探讨不同方法学对结直肠癌K-ras基因点突变检测结果的影响。[方法]对60例结直肠癌患者石蜡包埋组织样本进行K-ras基因第12、13位密码子突变检测,检测分别采用焦磷酸测序法及双脱氧测序法,并对结果进行对比分析。[结果]两种方法都能够检测到常见的突变类型,但由于两种方法的灵敏度不同,检出率焦磷酸测序法高于双脱氧测序,分别为33.3%(20/60)和25%(15/60)。经病理对肿瘤细胞比例分析发现,5例双脱氧测序法漏检标本中,肿瘤细胞比较少,不足20%。[结论]K-ras基因突变的检测过程中,标本肿瘤细胞的比例是影响结果的关键因素,检测前需要对标本进行病理评估,确保肿瘤细胞比例在50%以上。
[Objective] To investigate the influence of different methodologies on the detection of K-ras gene mutation in colorectal cancer. [Method] The codon 12 and 13 codons of K-ras gene were detected in paraffin-embedded tissue samples of 60 patients with colorectal cancer. Pyrosequencing and dideoxy sequencing were used respectively to detect the mutations in K-ras gene. The results were compared and analyzed. [Results] The common mutation types were detected by either method. However, due to the different sensitivities of the two methods, the detection rate of pyrosequencing was higher than that of dideoxy sequencing (33.3% (20/60) and 25% ( 15/60). The proportion of tumor cells by pathological analysis found that 5 cases of dideoxy sequencing missed samples, the tumor cells less, less than 20%. [Conclusion] The proportion of tumor cells in the detection process of K-ras gene mutation is the key factor affecting the results. Pathological evaluation of the specimens is required before the test to ensure that the proportion of tumor cells is above 50%.