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首页> 外文期刊>Molecular cytogenetics >Comprehensive chronic lymphocytic leukemia diagnostics by combined multiplex ligation dependent probe amplification (MLPA) and interphase fluorescence in situ hybridization (iFISH)
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Comprehensive chronic lymphocytic leukemia diagnostics by combined multiplex ligation dependent probe amplification (MLPA) and interphase fluorescence in situ hybridization (iFISH)

机译：通过结合多重连接依赖探针扩增（MLPA）和相间荧光原位杂交（iFISH）进行全面的慢性淋巴细胞白血病诊断

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Background Banding-karyotyping and metaphase-directed-fluorescence-in-situhybridization (FISH) may be hampered by low mitotic index in leukemia. Interphase FISH (iFISH) is a way out here, however, testing many probes at the same time is protracted and expensive. Here multiplex-ligation-dependent-probe-amplification (MLPA) was used retrospectively in chronic lymphocytic leukemia (CLL) samples initially studied by banding cytogenetics and iFISH. Detection rates of iFISH and MLPA were compared and thus a cost-efficient scheme for routine diagnostics is proposed. Results Banding cytogenetics was done successfully in 67/85 samples. DNA was extracted from all 85 CLL samples. A commercially available MLPA probe set directed against 37 loci prone to be affected in hematological malignancies was applied. Besides, routine iFISH was done by commercially available probes for following regions: 11q22.3, 12p11.2-q11.1, 13q14.3, 13q34, 14q32.33 and 17p13.1. MLPA results were substantiated by iFISH using corresponding locus-specific probes. Aberrations were detected in 67 of 85 samples (~79%) applying banding cytogenetics, iFISH and MLPA. A maximum of 8 aberrations was detected per sample; however, one aberration per sample was found most frequently. Overall 163 aberrations were identified. 15 of those (~9%) were exclusively detected by banding cytogenetics, 95 were found by MLPA (~58%) and 100 (~61%) by routine iFISH. MLPA was not able to distinguish reliably between mono- and biallelic del(13)(q14.3q14.3), which could be easily identified as well as quantified by routine iFISH. Also iFISH was superior to MLPA in samples with low tumor cell load. On the other hand MLPA detected additional aberrations in 22 samples, two of them being without any findings after routine iFISH. Conclusions Both MLPA and routine iFISH have comparable detection rates for aberrations being typically present in CLL. As MLPA can detect also rare chromosomal aberrations it should be used as an initial test if routine cytogenetics is not possible or non-informative. Still iFISH should be used additionally to distinguish mono- from biallelic deletions and also to determine rate of mosaicism for 13q14.2 to 13q14.3. In case MLPA is negative the corresponding CLL samples should be tested at least by iFISH using the standard probe set to.
机译：背景白血病中的低有丝分裂指数可能会阻碍带状核型分析和中期定向荧光原位杂交（FISH）。相间FISH（iFISH）是解决此问题的一种方法，但是，同时测试许多探针既耗时又昂贵。在这里，多重连接依赖性探针扩增（MLPA）被用于慢性淋巴细胞性白血病（CLL）样本的回顾性研究，该样本最初是通过结合细胞遗传学和iFISH研究的。比较了iFISH和MLPA的检出率，因此提出了一种具有成本效益的常规诊断方案。结果在67/85个样品中成功完成了带状细胞遗传学分析。从所有85个CLL样品中提取DNA。使用针对37种易受血液系统恶性肿瘤影响的基因座的市售MLPA探针组。此外，常规iFISH是通过市售探针针对以下区域进行的：11q22.3、12p11.2-q11.1、13q14.3、13q34、14q32.33和17p13.1。使用相应的基因座特异性探针通过iFISH证实了MLPA结果。使用条带细胞遗传学，iFISH和MLPA在85个样本中的67个（〜79％）中检测到畸变。每个样品最多检测到8个像差;但是，每个样本发现一个像差的频率最高。总共识别出163个像差。其中的15个（〜9％）仅通过条带细胞遗传学检测，MLPA发现了95个（〜58％），常规iFISH发现了100个（〜61％）。 MLPA无法可靠地区分单等位基因和双等位基因del（13）（q14.3q14.3），可以通过常规iFISH轻松识别和定量。在肿瘤细胞负荷低的样品中，iFISH也优于MLPA。另一方面，MLPA在22个样本中检测到其他像差，其中两个在常规iFISH后没有任何发现。结论MLPA和常规iFISH均具有可比较的CLL中常见畸变检出率。由于MLPA还可以检测罕见的染色体畸变，因此如果无法进行常规细胞遗传学研究或不能提供信息，则应将其用作初始检测。仍然应该另外使用iFISH来区分单等位基因缺失和双等位基因缺失，并确定13q14.2至13q14.3的镶嵌率。如果MLPA为阴性，则至少应使用设置为的标准探针通过iFISH测试相应的CLL样品。

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