微核、染色体试验用乙醇固定淋巴细胞可行性研究  

Feasibility study of lymphocytes being fixed by ethanol in micronucleus / chromosome test

作　　者：李茂进[1] 李春素[2] 赵培青 

机构地区：[1]北京协和医学院,北京100730 [2]北京市疾病预防控制中心,100013 [3]北京国济中医医院,100055

出　　处：《中国职业医学》2015年第4期417-420,424,共5页China Occupational Medicine

摘　　要：目的探讨微核、染色体试验中用乙醇部分代替甲醇固定淋巴细胞的可行性及相关技术方法。方法采用周围血半微量全血常规培养法进行试验。试验按固定液中甲醇和乙醇的比例不同分为Ⅰ、Ⅱ、Ⅲ、Ⅳ组,4种方法固定液中甲醇、乙醇和冰醋酸的比例分别为3∶0∶1、2∶1∶1、1∶2∶1、0∶3∶1(V∶V∶V),其他条件和操作均相同。取周围静脉血适量接种于RPMI-1640培养基中,于37℃培养72 h(染色体培养48 h),用体积分数为10%吉姆萨染液染色。结果Ⅰ、Ⅱ和Ⅲ组淋巴细胞均分散良好、染色均匀适中,胞浆呈蓝灰色,胞核呈紫红色,胞核轮廓清楚、边界清晰,均适合微核观察计数;Ⅳ组细胞容易聚集成团、胞浆易于破裂,致使适合观察的淋巴细胞数太少,从而影响微核的观察计数。Ⅰ、Ⅱ和Ⅲ组之间淋巴细胞着色良好率比较,差异有统计学意义(P<0.05);其中Ⅱ组淋巴细胞着色良好率高于Ⅲ组(P<0.05)。Ⅰ、Ⅱ和Ⅲ组淋巴细胞胞浆完整率和淋巴细胞微核细胞率分别比较,差异均无统计学意义(P>0.05)。Ⅰ、Ⅱ、Ⅲ和Ⅳ组淋巴细胞细胞成团指数比较,差异有统计学意义(P<0.05);其中Ⅰ、Ⅱ和Ⅲ组细胞成团指数均低于Ⅳ组(P<0.05)。Ⅰ、Ⅱ和Ⅲ组淋巴细胞染色体呈紫红色、分散良好、大小适中、背景清楚,均适合染色体观察;Ⅳ组不适用于染色体观察。结论微核和染色体试验的固定液中加入一定比例的乙醇(乙醇与甲醇的体积比不超过2∶1)代替甲醇固定淋巴细胞,不影响微核和染色体的观察,并可降低甲醇中毒风险。Objective To explore the feasibility and relevant technology of using ethanol as a partial substitute for methanol to fix lymphocytes in micronucleus and chromosome test. Methods The micro-whole blood routine culture method was used to determine the micronuleus and chromosome of lymphocytes in peripheral blood. According to the ratio of methanol to ethanol in fixing solutions, 4 groups of I , Ⅱ, Ⅲ and IV were set up, in which the ratios of methanol, ethanol, and glacial acetic acid （ V： V： V） were 3： 0： 1, 2： 1： 1, 1： 2： 1, and 0： 3： 1, respectively. The test was conducted under the same operations and conditions. The lymphocytes from the peripheral venous blood were cultured in RPMI-1640 at 37 ~C for 72 hours （48 hours for chromosomes） ; dyed with 10% Giemsa （ V： V）. Results The lymphocytes in peripheral blood of group I , Ⅱ , and m were all well-distributed and stained purely, full of blue gray in cytoplasm, and the nuclei were purple red with clear outline and boundary, which completely met the observation and counting of micronucleus. The lymphocytes in peripheral blood of group IV tended to cluster and the cytoplasm was easy to rupture, resulting in less numbers of lymphocytes suitable for observation, so as to influence the observation of micronucleus counting. Differences were statistically significant among the good staining rates of lymphocytes in groups of i , Ⅱ , and Ⅲ （P 〈 0.05 ）, and the good staining rate of group Ⅱ was higher than that of group m （ P 〈 0. 05 ）. The lymphocyte cytoplasm integrity rates and micronuclei frequency of lymphocytes of the 3 groups of I , Ⅱ and Ⅲ showed no significant difference （ P 〉 0. 05 ）. The lymphocyte cluster indices of the 4 groups showed statistical significant difference （ P 〈 0. 05）, and the lymphocyte cluster index of group IV had significant higher compared with those of the other 3 groups （ P 〈 0.05）, not suitable for the observation of chromosomes and micronuclei. The chromoso

关 键 词：淋巴细胞 微核 染色体 乙醇 甲醇 试验方法 

分 类 号：R135.1[医药卫生—劳动卫生] R995[医药卫生—公共卫生与预防医学]