Quantum dots thermal stability improves simultaneous phenotype-specific telomere length measurement by FISH-flow cytometry

In this study, we have substituted quantum dots (nanoparticles) for traditional fluorophores in FISH-flow. Quantum dots were demonstrated to possess much greater thermal stability than traditional low molecular weight and phycobiliprotein fluorophores. Quantum dot antibody conjugates directed against monocyte and T cell antigens were found to retain most of their fluorescence following the high temperature annealing step, allowing simultaneous fluorescent immunophenotyping and telomere length measurement. Since quantum dots have very narrow emission bandwidths, we were able to analyze multiple quantum dot antibody conjugates (Qdot 605, 655 and 705) simultaneously with FISH-flow measurement to assess the age-associated decline in telomere length in both human monocytes and T cell subsets. With quantum dot immunolabeling, the mean decrease rate in telomere length for CD4+ cells was calculated at 41.8 bp/year, very close to previously reported values using traditional flow-FISH and Southern blotting. This modification to the traditional flow-FISH technique should therefore allow simultaneous fluorescent immunophenotyping and telomere length measurement, permitting complex cell subset-specific analysis in small numbers of cells without the requirement for prior cell sorting.