Gole Leena, Lin Adeline, Chua Constance, Chng Wee Joo
Department of Laboratory Medicine, National University Health Systems, Singapore.
Department of Haematology Oncology, National Cancer Institute of Singapore, Singapore.
Cancer Genet. 2014 Jan-Feb;207(1-2):31-4. doi: 10.1016/j.cancergen.2013.12.001. Epub 2013 Dec 27.
The International Myeloma Working Group recommends that fluorescence in situ hybridization (FISH) be performed on specifically identified plasma cells (PC). This is because chromosomal abnormalities are not frequently detected by traditional karyotyping due to the low proliferative rate of PC in multiple myeloma (MM). Conventional FISH enhances the sensitivity but lacks the specificity, as it does not distinguish PC from other hematopoetic cells. To fulfill this recommendation, PC need to be selected either by flow cytometry or immunomagnetic bead-based PC sorting or by concomitant labeling of the cytoplasmic immunoglobulin light chain, which allows for unambiguous identification. These techniques require expertise, time, and funding and are not easily incorporated into the routine workflow of the cytogenetic laboratory. We have modified and refined the technique using fixed cell pellets to achieve nicely separated and easily identifiable PC. With immunostaining and subsequent FISH (i.e., cytoplasmic immunoglobulin FISH, cIg-FISH), this technique can be easily incorporated into every cytogenetic laboratory. Twenty samples from patients with MM were subjected to routine FISH, cIg-FISH, and chromosomal karyotyping and the results were compared. Three FISH probes, which enabled detection of the t(4;14), t(14;16) and deletion of TP53, were used to validate this modified technique successfully.
国际骨髓瘤工作组建议对经特异性鉴定的浆细胞（PC）进行荧光原位杂交（FISH）检测。这是因为在多发性骨髓瘤（MM）中，由于浆细胞的增殖率较低，传统的核型分析并不经常能检测到染色体异常。传统的FISH提高了敏感性，但缺乏特异性，因为它无法将浆细胞与其他造血细胞区分开来。为了满足这一建议，需要通过流式细胞术或基于免疫磁珠的浆细胞分选，或者通过同时标记细胞质免疫球蛋白轻链来选择浆细胞，这样才能明确识别。这些技术需要专业知识、时间和资金，并且不容易纳入细胞遗传学实验室的常规工作流程。我们对使用固定细胞沉淀的技术进行了改进和完善，以实现浆细胞的良好分离和易于识别。通过免疫染色和随后的FISH（即细胞质免疫球蛋白FISH，cIg-FISH），该技术可以很容易地纳入每个细胞遗传学实验室。对20例MM患者的样本进行常规FISH、cIg-FISH和染色体核型分析，并比较结果。使用三种能够检测t(4;14)、t(14;16)和TP53缺失的FISH探针成功验证了这种改良技术。
