Quantifying telomere lengths of human individual chromosome arms by centromere-calibrated fluorescence in situ hybridization and digital imaging.

Telomere length analysis has aroused considerable interest in biology and oncology. However, most published data are pan-genomic Southern-blot-based estimates. We developed T/C-FISH (telomere/centromere-FISH), allowing precise measurement of individual telomeres at every single chromosome arm. Metaphase preparations are co-hybridized with peptide nucleic acid probes for telomeric sequences and the chromosome 2 centromere serving as internal reference. Metaphase images are captured and karyotyped using dedicated software. A software module determines the absolute integrated fluorescence intensities of the p- and q-telomeres of each chromosome and the reference signal. Normalized data are derived by calculating the ratio of absolute telomere and reference signal intensities, and descriptive statistics are calculated. T/C-FISH detects even small differences in telomere length. Using T/C-FISH we have discovered an epigenetic process occurring in the human male postzygote or early embryo: in umbilical cord blood lymphocytes, telomeres on male Xqs are around 1100 bp shorter than female Xqs.