Cell Analysis – Cell Proliferation Imaging Assays

Cell analysis allows scientists to gain unique insights into cell proliferation, viability, and toxicity, enabling the monitoring of cell response and cell health in cultures after treatment with various stimuli.

Detection of DNA replication is a powerful tool for studying cell proliferation of both cell populations and individual cells. DNA replication can be detected through the incorporation of a nucleoside analog such as BrdU or EdU into DNA. The detection of these two thymidine analogs differs significantly and has implications for test results, potential applications, and multiplexing capability. Compared to the other cell proliferation methods, the EdU proliferation assay does not involve radioactive isotopes or antibodies to detect the newly synthesized DNA.

The ClickTech EdU cell proliferation assay enables estimates of cell proliferation rates by determining the fraction of cells replicating their DNA.The technology is based on the labeling of DNA with EdU (5-ethynyl-2′-deoxyuridine), an alkyne-modified nucleoside that is incorporated into the replicating DNA of living cells and its conjugation to a fluorochrome in situ.

Schematic representation of EdU cell proliferation assays. A) Incubation of cells with EdU. B) EdU is incorporated during active DNA synthesis. C) Detection of cell proliferation via click chemistry, wherein the use of a range of different fluorescent dyes is possible.

Advantages:

• Increased selectivity and sensibility in comparison to BrdU
• Simple, fast and reliable assay
• High sensitivity paired with very low toxicity
• A cost-effective assay
• Multiplexing possibility
• higher preservation of morphological details relative to BrdU

ClickTech EdU Cell Proliferation Assays for Imaging

The baseclick ClickTech EdU cell proliferation assays overcome the limitations of conventional cell proliferation assays and offer a superior alternative to BrdU and [3H] thymidine assays. EdU (5-ethynyl-2′-deoxyuridine) is a thymidine analog, which is incorporated into DNA during active DNA synthesis. The optimized detection protocol considerably simplifies the workflow and the time required. The simple detection procedure is completed within 30 minutes and is compatible with multiplexing for content- and context-rich results.

Workflow

EdU Detection

Fluorescence imaging of replicating HeLa cells detected with the EdU Cell Proliferation Kit for Imaging (BCK-EdU488IM100). Green signal: EdU labelling; blue signal: DNA counterstaining with DAPI; scale bar = 10 µm. Distinct replication patterns allow staging of replicating cells along S phase (early/mid/late S phase).

Cell proliferation assay with the EdU Cell Proliferation Kit for Imaging (BCK-EdU647IM100). A) Representative fluorescence microscopy image. Red signal: EdU labelling; blue signal: DNA counterstaining with DAPI. B) Cells in S, G1 and G2/M phases are quantified by image cytometry.

Product Overview

LITERATURE

Anticancer drug and ionizing radiation-induced DNA damage differently influences transcription activity and DDR-related stress responses of an endothelial monolayer, V. Ziegler et al., 2020, Molecular Cell Research, Vol. 1867, p. 118678.

https://doi.org/10.1016/j.bbamcr.2020.118678

Dendrimer-Based Signal Amplification of Click-Labelled DNA in Situ, N. Raddaoui et al. 2017, ChemBioChem, Vol. 18, p. 1716-1720.

https://doi.org/10.1002/cbic.201700209

High-copy sequences reveal distinct evolution of the rye B chromosome, S. Klemme et al., 2013, New Phytologist, Vol. 199, p. 550-558.

https://doi.org/10.1111/nph.12289

Cytolethal Distending Toxin Promotes Replicative Stress Leading to Genetic Instability Transmitted to Daughter Cells, W. Tremblay et al., 2021, Frontiers in Cell and Developmental Biology, Vol. 9, 656795.

https://doi.org/10.3389/fcell.2021.656795

Brassica napus Roots Use Different Strategies to Respond to Warm Temperatures, M. Boter et al., 2023, Int. J. Mol. Sci., Vol. 24(2), 1143.

https://doi.org/10.3390/ijms24021143