Ege University, School of Medicine, Medical Biology Department, İzmir, Turkey.
Ege University, School of Medicine, Pathology Department, İzmir, Turkey.
Turk J Haematol. 2012 Jun;29(2):126-34. doi: 10.5505/tjh.2012.93898. Epub 2012 Jun 15.
Follicular lymphoma (FL) is one of the most common lymphomas, and is characterized by t(14;18)(q32;q21) in more than 80% of patients. The aim of this study was to determine the rate of t(14;18) positivity based onthe detection of mbr or mcr in paraffin-embedded tissue samples.
The study included 32 paraffin-embedded tissue samples collected from 32 consecutive FL patients that were diagnosed and followed-up at our hospital between 1999 and 2006. The MBR breakpoint wasidentified based on real-time PCR using a LightCycler v.2.0 t(14;18) Quantification Kit (MBR), multiplex PCR, and seminestedPCR. To identify the mcr breakpoint, real-time PCR was performed using specific primers and the FastStart DNAMaster SYBR Green I Kit. To detect t(14;18) via fluorescence in situ hybridization (FISH) nuclei from paraffin-embeddedtissue sections were extracted and used together with LSI IgH (immunoglobulin heavy chain) (spectrum green)/bcl-2(B-cell leukemia-lymphoma 2) (spectrum orange) probes.
The DNA and nuclei isolation success rate for B5 formalin-fixed, paraffin-embedded tissue sections (n = 12)was 42% and 33%, respectively, versus 95% and 60%, respectively, for 20 tissue sections fixed in formalin only. In all,24 paraffin-embedded tissue sections were analyzed and mbr positivity was observed in the DNA of 82.14% via seminested PCR, in 53.57% via multiplex PCR, and in 28.57% via real-time PCR. We did not detect mcr rearrangementin any of the samples. In all, 15 of 16 patients (93.75%) whose nuclei were successfully isolated were observed to bet(14;18) positive via the FISH method.
Semi-nested PCR and FISH facilitated the genetic characterization of FL tumors. As such, FISH and PCR complement each other and are both essential for detecting t(14;18) translocation.
滤泡性淋巴瘤（FL）是最常见的淋巴瘤之一，其特征在于超过 80%的患者存在 t(14;18)(q32;q21)。本研究的目的是基于石蜡包埋组织样本中 mbr 或 mcr 的检测来确定 t(14;18) 的阳性率。
本研究纳入了 32 例石蜡包埋组织样本，这些样本来自于 1999 年至 2006 年间在我院确诊和随访的 32 例连续 FL 患者。使用实时 PCR 通过 LightCycler v.2.0 t(14;18) Quantification Kit（MBR）、多重 PCR 和半巢式 PCR 来确定 MBR 断点。为了确定 mcr 断点，使用特定引物和 FastStart DNA Master SYBR Green I Kit 进行实时 PCR。通过荧光原位杂交（FISH）检测 t(14;18)，从石蜡包埋组织切片中提取核，并与 LSI IgH（免疫球蛋白重链）（谱绿色）/bcl-2（B 细胞白血病-淋巴瘤 2）（谱橙色）探针一起使用。
B5 福尔马林固定、石蜡包埋组织切片的 DNA 和核分离成功率分别为 42%和 33%，而仅用福尔马林固定的 20 个组织切片的成功率分别为 95%和 60%。总共分析了 24 个石蜡包埋组织切片，通过半巢式 PCR 在 DNA 中观察到 mbr 阳性率为 82.14%，通过多重 PCR 为 53.57%，通过实时 PCR 为 28.57%。我们没有在任何样本中检测到 mcr 重排。在所有成功分离核的 16 例患者（93.75%）中，通过 FISH 方法观察到 15 例为 t(14;18)阳性。
半巢式 PCR 和 FISH 促进了 FL 肿瘤的遗传特征分析。因此，FISH 和 PCR 相互补充，都是检测 t(14;18)易位所必需的。
