Martens U M, Brass V, Engelhardt M, Glaser S, Waller C F, Lange W, Schmoor C, Poon S S, Landsdorp P M
Freiburg University Medical Center, Department of Hematology/Oncology, Germany.
Biochem Soc Trans. 2000 Feb;28(2):245-50. doi: 10.1042/bst0280245.
The DNA of human chromosomes terminates in several kilobases of telomere repeats that are gradually lost with; age and with replication in vitro. Defective telomere maintenance has been shown to be causally linked to cell cycle exit and apoptosis. In order to overcome the limitations imposed by Southern blotting, we have established a quantitative fluorescence in situ hybridization (Q-FISH) technique. This technique allows estimation of telomere length in specific chromosome arms from metaphase cell preparations. Furthermore, we have extended quantitative in situ hybridization to flow cytometry (flow FISH) in order to obtain information on the mean telomere repeat content in suspended cells. Telomere length in granulocytes, monocytes, CD8 and CD4 T lymphocytes and natural killer cells was found to differ slightly in the peripheral blood of adults. However, strikingly longer telomeres were observed in B lymphocytes (approximately 1.3 kb longer), suggesting a functional role for telomere maintenance in this cell subset. In summary, Q-FISH and flow FISH represent new methods for measuring telomere length in single cells and allow studies of telomere dynamics in haematopoietic subpopulations at various stages of normal and abnormal antigen responses.
人类染色体的DNA末端是几千个碱基对的端粒重复序列，这些序列会随着年龄增长以及体外复制而逐渐丢失。已证明端粒维持缺陷与细胞周期退出和细胞凋亡存在因果关系。为了克服Southern印迹法的局限性，我们建立了一种定量荧光原位杂交（Q-FISH）技术。该技术可从有丝分裂中期细胞制备物中估计特定染色体臂的端粒长度。此外，我们已将定量原位杂交扩展到流式细胞术（流式FISH），以便获取悬浮细胞中端粒重复序列平均含量的信息。在成年人外周血中发现，粒细胞、单核细胞、CD8和CD4 T淋巴细胞以及自然杀伤细胞的端粒长度略有不同。然而，在B淋巴细胞中观察到端粒明显更长（约长1.3 kb），这表明端粒维持在该细胞亚群中具有功能性作用。总之，Q-FISH和流式FISH代表了测量单细胞中端粒长度的新方法，并允许研究正常和异常抗原反应各个阶段造血亚群中的端粒动态。
