Beliën J A, Copper M P, Braakhuis B J, Snow G B, Baak J P
Department of Pathology, Free University Hospital, Amsterdam, The Netherlands.
Carcinogenesis. 1995 Oct;16(10):2395-400. doi: 10.1093/carcin/16.10.2395.
The proportion of exfoliated buccal mucosal cells with micronuclei gives the opportunity to assess sensitivity to gamma-radiation and genotoxic compounds and in addition to monitor the effectiveness of cancer intervention strategies. So far, results on counting micronuclei in various publications are difficult to compare because of differences in methods used, especially with regard to microscopical magnification used and number of cells counted. The aims of this study were (i) to define a protocol for counting micronuclei; (ii) to assess the feasibility of manually counting micronuclei; and (iii) the assessment of inter- and intra-patient variability of the number of micronuclei. We propose the definition of a strict protocol on counting micronuclei, with regard to cytological preparation, definition of micronuclei, instrumentation, sampling of cells in a cytological specimen and sample size. Such a strict protocol is a prerequisite for counting micronuclei in exfoliated cells to get a reproducible and sensitive indicator of exposure and for cancer risk. Although the inter- and intra-observer reproducibility of counting micronuclei per 1000 cells using such a protocol is well, we show that the variability among 10 assessments of micronuclei per 1000 cells taken sequentially from a sample size of 10,000 nuclei of the same specimen can be enormous (coefficients of variation varied in seven individuals studied between 42.1 and 102.9%). Based on the observed low frequencies varying from 1.2 to 5.2 micronuclei per 1000 cells and the variation found, we conclude that at least 10,000 exfoliated cells should be screened to monitor a significant reduction of 50% in the number of micronuclei (for a patient with an initial frequency in the micronuclei frequency range given). Since it takes approximately 7 h to evaluate this number of cells, it is also concluded that counting of micronuclei requires automation.
具有微核的脱落颊黏膜细胞比例为评估对γ辐射和遗传毒性化合物的敏感性提供了机会，此外还可监测癌症干预策略的有效性。到目前为止，由于所使用方法的差异，尤其是在显微镜放大倍数和计数细胞数量方面，不同出版物中微核计数的结果难以比较。本研究的目的是：（i）定义微核计数方案；（ii）评估手动计数微核的可行性；（iii）评估患者间和患者内微核数量的变异性。我们针对细胞制备、微核定义、仪器设备、细胞学标本中的细胞采样和样本量，提出了严格的微核计数方案定义。这样一个严格的方案是在脱落细胞中计数微核以获得可重复且敏感的暴露指标以及癌症风险指标的先决条件。尽管使用该方案每1000个细胞计数微核的观察者间和观察者内可重复性良好，但我们表明，从同一样本的10000个细胞核中依次抽取的10次每1000个细胞微核评估之间的变异性可能非常大（在7名研究个体中，变异系数在42.1%至102.9%之间变化）。基于观察到的每1000个细胞中微核频率较低，在1.2至5.2之间，以及所发现的变异性，我们得出结论，至少应筛选10000个脱落细胞，以监测微核数量显著减少50%（对于微核频率在给定范围内的初始患者）。由于评估这个细胞数量大约需要7小时，因此还得出结论，微核计数需要自动化。
