CyTOF Mass Cytometry for Click Proliferation Assays

Area of Science:

• Cell Biology
• Biochemistry
• Immunology

Background:

• Novel cell analyzers like polychromatic flow cytometers and mass cytometers (CyTOF) allow extensive single-cell analysis.
• Traditional BrdU assays for cell proliferation have limitations, including DNA denaturation and inability to resolve complex phenotypes.
• Click chemistry offers a solution to the limitations of BrdU assays.

Purpose of the Study:

• To introduce a novel assay for active S-phase determination in cell cycle analysis.
• To combine click chemistry with iodo-deoxyuridine (IdU) for detecting DNA synthesis.
• To provide a method compatible with both flow cytometry and CyTOF analysis.

Main Methods:

• Utilized click chemistry with a novel IdU azide derivative.
• Employed a cross-reactive anti-IdU antibody to detect incorporated ethynyl-labeled nucleotides (EdU).
• Developed a protocol bridging flow cytometry and CyTOF for cell cycle analysis.

Main Results:

• The novel assay successfully detects incorporated EdU during DNA synthesis.
• The method is suitable for complex and multiparametric experiments.
• It provides a cost-effective approach for ethynyl-labeled nucleotide detection.

Conclusions:

• The IdU click chemistry assay is a preferred alternative to BrdU assays for cell cycle analysis.
• This method facilitates simultaneous flow and mass cytometry analyses.
• It offers a feasible and advantageous approach for advanced cell proliferation studies.