Flow cytometric measurement of absolute telomere length

Abstract

The length of the end of chromosomes – telomeres is a dynamic and constant characterizes cellular aging process. Measurement of telomere length (TL) and its connection with the diagnosis, course, prognosis and treatment of a number of diseases in recent years, is of great interest for researchers. The methods used for this are varied and are primarily by molecular biological. Variability of methodological approaches, the lack of a common standard of measurement TL measurements in absolute and relative terms: all this leads to results that cannot be compared with each other. The aim of this work is the development of data conversion algorithm is obtained by flow cytometry in absolute telomere length, expressed in kilobases (kb). The study was conducted from venous blood of children 2–3 years of age (8 healthy children and 3 patients with dyskeratosis congenita). As control material used cell line 1301 (ATCC, UK). Determination of relative telomere length was performed by using a set FlowFISH Telomere PNA Kit / FITC (DakoCytomation, Glostrup, Denmark). RTL code and the absolute length of telomeres were calculated for each study participant. Data on the length of patients telomere were compared with data on TL healthy children of appropriate age. RTL average value for healthy children 2–3 years was 30.1, and an absolute telomere length – 15.56 kb. Telomere length patients with dyskeratosis congenita was significantly shorter (RTL 1.4–4.3, the absolute value of TL 0.3–2.4 kb). The algorithm conversion molecular fluorescence equivalent in absolute TL is universal and can be used in any laboratory equipped with a flow cytometer. The resulting data TL can be compared with the data expressed in absolute units and obtained by any other method.