Validation and development of quantitative flow cytometry-based fluorescence in situ hybridization for intercenter comparison of telomere length measurement.
BACKGROUND
Telomeres are highly conserved repeats at the ends of chromosomes that maintain chromosome stability and reflect the replicative potential of cells. Telomere length can be determined by Southern blot hybridization or quantitative fluorescence in situ hybridization (Q-FISH). Recently, two flow cytometry-based (Flow) FISH protocols have been published.


METHODS
We compared the telomere length measured by Southern blotting and Flow FISH using standard beads to calibrate and quantify the fluorescence intensity.


RESULTS
The telomeric fluorescence of cord blood and peripheral blood mononuclear cells was similar to that reported by other studies. There was a linear relationship between the telomeric fluorescence determined by Flow FISH and the telomere fragment size determined by Southern blotting (r = 0.89; P < 0.001).


CONCLUSIONS
It is important to set up a center-specific curve and select appropriate cell lines for reference. This Q-Flow FISH protocol will facilitate the measurement of telomere length and allow more meaningful comparison of data (in standard fluorescence units or fragment size) between institutes.
Law
H
H
Department of Paediatrics, Faculty of Medicine, The University of Hong Kong, Queen Mary Hospital, Pokfulam, Hong Kong.
Lau
Y
Y
D000328:Adult; D015139:Blotting, Southern / Q000706:statistics & numerical data; D002138:Calibration; D004273:DNA, Neoplasm / Q000032:analysis; D005312:Fetal Blood / Q000737:chemistry / Q000166:cytology; D005434:Flow Cytometry / Q000379:methods* / Q000706:statistics & numerical data; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods* / Q000706:statistics & numerical data; D019169:Jurkat Cells; D020014:K562 Cells; D008875:Middle Aged; D015203:Reproducibility of Results; D015183:Restriction Mapping / Q000379:methods; D016615:Telomere / Q000737:chemistry*
D004273:DNA, Neoplasm