Elenitoba-Johnson Kojo S J
Division of Anatomic Pathology, University of Utah Health Sciences Center, Salt Lake City, Utah 84132, USA.
Methods Enzymol. 2002;356:224-40. doi: 10.1016/s0076-6879(02)56936-6.
DNA sequencing of the antigen receptor genes remains the gold standard for the establishment of clonal relationships between samples. However, a variety of strategies may be employed as surrogates for the determination of the actual sequence of the clonally rearranged antigen receptor genes. The methods described in this chapter provide a framework for the rapid determination of clonal relationships between (microdissected) lymphoid populations. All of the methods described are PCR-based because of its versatility and ability to utilize very small amounts of DNA. For illustration purposes, the descriptions have been confined to B-cell populations. Although not described here, Ig kappa or Ig lambda PCR may also be utilized for determination of B-cell clonality and clonal relationships in the same manner. Similarly, the principles utilized may be extended to T-cell populations and T-cell receptor chain genes. Regardless of the methodology or targets involved, it is strongly recommended that all assays on microdissected material be run on parallel replicates of each sample to ensure reproducibility of results. The information about clonal relationships obtained by LCM has more than an academic significance and has utility in routine diagnostics for the establishment of minimal residual disease and the determination of microscopic disease recurrence vs the development of a secondary malignancy.
抗原受体基因的DNA测序仍然是确定样本间克隆关系的金标准。然而，可采用多种策略作为替代方法来确定克隆重排的抗原受体基因的实际序列。本章所述方法为快速确定（显微切割的）淋巴样群体间的克隆关系提供了一个框架。由于其通用性以及能够利用极少量DNA的能力，所述所有方法均基于PCR。为便于说明，描述仅限于B细胞群体。虽然此处未作描述，但Ig κ或Ig λ PCR也可同样用于确定B细胞的克隆性和克隆关系。同样，所采用的原理可扩展至T细胞群体和T细胞受体链基因。无论涉及何种方法或靶点，强烈建议对显微切割材料进行的所有检测都在每个样本的平行重复样本上进行，以确保结果的可重复性。通过激光捕获显微切割获得的克隆关系信息不仅具有学术意义，在常规诊断中对于确定微小残留病以及区分显微镜下疾病复发与继发性恶性肿瘤的发生也具有实用价值。
