A novel method for kinetic measurements of rare cell proliferation using Cellometer image-based cytometry.

Cell proliferation is an important assay for pharmaceutical and biomedical research to test the effects of a variety of treatments on cultured primary cells or cell lines. For immunological studies, the ability to perform rapid cell proliferation analysis allows the identification of potential biological reagents for inducing or inhibiting immune cell proliferation. Current cell proliferation analysis methods employ flow cytometry for fluorescence detection of CFSE-labeled cells. However, conventional flow cytometers require a considerable amount of cells per sample, which becomes an issue for kinetic measurements with rare cell population due to the lack of samples for flow cytometric analyses at multiple time points during proliferation period. Here we report the development of a novel cell proliferation kinetic detection method for low cell concentration samples using the new Cellometer Vision system. Since the Cellometer system requires only 20 μl of sample, cell proliferation can be measured at multiple time points over the entire culturing period, whereas typically, flow cytometry is only performed at the end of the proliferation period. To validate the detection method, B1 and B2 B cells were treated with a B cell mitogen for 6 days, and proliferation was measured using Cellometer on day 1, 3, 5, and 6. To demonstrate the capability of the system, B1 B cells were treated with a panel of TLR agonists (Pam3Cys, PolyIC, CLO97, and CpG) for 7 days, and proliferation was measured on day 2, 4, 6, and 7. Cellometer image-based cytometry (IBC) was able to obtain proliferation results on each day with the last time point comparable to flow cytometry. This novel method allows for kinetic measurements of the rare cell samples such as B1 B cell, which has the potential to revolutionize kinetic analysis of cell proliferation.