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Journal of ZheJiang University(Medical Science)  2017, Vol. 46 Issue (4): 439-444    DOI: 10.3785/j.issn.1008-9292.2017.08.15
    
Volume of 4% neutral buffered formaldehyde affects the results of HER2 gene detected by fluorescence in situ hybridization in primary invasive breast cancer
CHEN Zhiqiang, WANG Ying, MI Xianjun, DUAN Lifeng, CHEN Ang, HUANG Huayong
Department of Pathology, Zhongshan Boai Hospital Affiliated to Southern Medical University, Zhongshan 528400, China
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Abstract  

Objective:To investigate the effects of the volume of 4% neutral phosphate buffered formalin fixative solution on the detection of human epidermal growth factor receptor 2 (HER2) gene by fluorescence in situ hybridization (FISH) in primary invasive breast cancer. Methods:Tissue samples were collected from 109 patients with primary invasive breast cancer admitted in Zhongshan Boai Hospital from June 2014 to October 2016. The ratios of 4% phosphate buffered formalin fixative solution to sample volume samples were 3:1, 6:1, 9:1, 10:1, 15:1, 20:1 or 25:1 (groups A, B, C, D, E, F and G), respectively. Paraffin sections were made after 15 h of fixation. The amplification of HER2 gene was detected by FISH. The gene amplification results of HER2 were observed and compared in different groups. Results:Fluorescence microscope showed that the tissue contour in groups A, B and C was vague, cell debris appeared, and the probe was positioned poorly; while the tissue contour was clear and complete in groups D, E, F and G and the probe was positioned accurately. The positive rate of HER2 was gradually increased from group A to D(χ2=8.601, P<0.01), and that remained stable at 24.77% in groups D to G. The positive rate of gene amplification in groups D, E, F and G was significantly higher than that in groups A, B and C (all P<0.05). Conclusion:When using FISH to detect HER2 gene in samples of primary breast invasive carcinoma, the volume of fixative solution should be at least 10 times of the sample volume to obtain accurate and stable results.



Key wordsBreast neoplasms/pathology      Neoplasm invasiveness      Fixatives      Formaldehyde/administration &      dosage      In situ hybridization, fluorescence      Receptor, epidermal growth factor     
Received: 17 March 2017      Published: 25 August 2017
CLC:  R446.6  
Cite this article:

CHEN Zhiqiang, WANG Ying, MI Xianjun, DUAN Lifeng, CHEN Ang, HUANG Huayong. Volume of 4% neutral buffered formaldehyde affects the results of HER2 gene detected by fluorescence in situ hybridization in primary invasive breast cancer. Journal of ZheJiang University(Medical Science), 2017, 46(4): 439-444.

URL:

http://www.zjujournals.com/med/10.3785/j.issn.1008-9292.2017.08.15     OR     http://www.zjujournals.com/med/Y2017/V46/I4/439


不同量甲醛固定液对荧光原位杂交法检测乳腺原发性浸润癌HER2基因扩增的影响

目的:通过分析不同的体积量甲醛固定液对荧光原位杂交(FISH)法检测乳腺原发性浸润癌人类表皮生长因子受体2(HER2)基因扩增的差异,为建立和优化乳腺癌HER2基因扩增检测过程中的标本固定方法提供实验依据。方法:选取中山市博爱医院2014年6月至2016年10月手术切除乳腺原发性浸润癌109例,同一肿瘤病变部位切取组织七块,根据随机数字表分为A、B、C、D、E、F、G组。使用4%中性(磷酸盐缓冲)甲醛作为固定液,从A组到G组固定液量与所浸泡组织体积的比例分别为3:1、6:1、9:1、10:1、15:1、20:1、25:1,固定15 h后制作石蜡切片。采用FISH法检测HER2基因扩增,观察各组HER2基因扩增结果的差异。结果:在荧光显微镜下,A、B、C组的组织轮廓较模糊,出现细胞碎片,部分探针定位欠佳,出现信号丢失、核发空现象,见明亮的桔红/绿色荧光信号;D、E、F、G组的组织轮廓清晰而完整、背景洁净、探针定位准确,见耀眼的桔红/绿色荧光信号。A~D组基因扩增阳性率逐渐增高(χ2=8.601,P<0.01),D~G组基因扩增阳性保持稳定;固定液量与所浸泡组织体积的比例在10:1以下(A、B、C组)时,HER2基因扩增状态不稳定,随着固定液量的增加不断出现新的基因扩增阳性患者;固定液量与所浸泡组织体积的比例在10:1及以上(D、E、F、G组)时,HER2基因扩增阳性率稳定在24.77%;D、E、F、G组的基因扩增阳性率高于A、B、C组(均P<0.05)。结论:采用FISH法检测乳腺原发性浸润癌HER2基因扩增,固定液的量要保证至少是所浸泡组织体积的10倍以上方可获得确切而稳定的实验结果。


关键词: 乳腺肿瘤/病理学,  肿瘤浸润,  固定剂,  甲醛/投药和剂量,  原位杂交,  荧光,  受体,  表皮生长因子 

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