苏木素复染对免疫组化显色强度定量分析的影响

Effect of hematoxylin counterstain on the quantitative immunohistochemical staining analysis

目的探讨苏木素复染是否影响免疫组化显色反应强度,为免疫组化定量分析的质量控制提供科学依据。方法用免疫组织化学方法检测GLC-82肺腺癌细胞株细胞蜡块中Ki67、Tiam1的表达,检测分为复染组和非复染组;在Nikon显微镜下通过Canon数码像机和Zoom Browser Ex采图软件随机截取样品显色反应细胞图像,用Imagepro Plus图像分析软件,分别测试苏木素复染与不复染时GLC-82细胞Tiam1、Ki67蛋白表达的阳性单位(PU值),每组各测试100个细胞。结果苏木素复染后GLC-82细胞Tiam1的PU值为(36.76±5.12),大于未复染Tiam1的PU值(26.64±5.09,t=9.906,P=0.000),相对偏差为38%。苏木素复染后GLC-82细胞Ki67的PU值为(21.74±4.59),大于未复染Ki67的PU值(14.37±4.65,t=7.985,P=0.000),相对偏差为51%。结论经苏木素复染,Tiam1、Ki67的PU值均明显增大,且核表达抗原Ki67的相对偏差明显大于浆表达抗原Tiam1的相对偏差。因此,对免疫组化结果定量分析时,为保证测试结果的可靠性,不应进行苏木素复染。

Objective To investigate whether the hematoxylin counterstain can affect the color intensity of cells after immunohistochemical staining,and to provide a basis for quality control of immunohistochemical staining.Methods Immunohistochemical staining method was used to detect the expression of Ki67 and Tiam1 in human lung adenocarcinoma cell line GLC-82.The samples were divided into counterstaining and non-counterstaining groups.Through the Zoom Browser Ex photograph System,the positive image was captured under the Nikon microscope equipped Canon digital camera.The positive unit（PU） of Ki67 and Tiam1 in GLC-82 cells with or without hematoxylin counterstaining were singly assessed quantitatively using Imagepro Plus image analysis software.Results The PU value of Tiam1 was greater in GLC-82 cells with hematoxylin counterstaining（36.76±5.12） than in those without hematoxylin counterstaining（26.64±5.09,t=9.906,P=0.000）,and the relative deviation was 38%.The PU value of Ki67 was greater in GLC-82 cells with hematoxylin counterstaining（21.74±4.59） than in those without hematoxylin counterstaining（14.37±4.65,t=7.985,P=0.000）,and the relative deviation was 51%.Conclusion PU values of Tiam1 and Ki67 were greatly increased in cells with hematoxylin counterstaining,and the relative deviation of Ki67 expression in nuclear was greater than Tiam1 in the cytoplasm.To ensure the result accuracy,hematoxylin counterstaining should not be used in tissue sections with immunohistochemical staining.