Performing a cell killing assay without using radioactive materials

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Have you ever thought of performing a cell killing assay without using radioactive materials?

Phenomics Australia node at the VCFG recently teamed up with Dr Kevin Sek to develop an application using their Incucyte SX5 Live Cell Analysis instrument.

Dr Sek tracked live tumour (adherent and non-adherent) cell growth over 48 hours in the presence or absence of CAR T cells. He was able to show that CAR T cells were effectively killing tumour cells over time. He captured images and videos of the whole process in 384-well plate format, allowing him to test several conditions in a single experiment. In comparison, if he performed the gold standard chromium killing assay, in a 96-well format, he would use substantially more cells, would be limited in the number of test conditions and would only have a single end point assay result.

This is such a great advance!

Dr Kevin Sek (R), Postdoctoral Researcher at the Peter MacCallum Cancer Centre and Dr Henry Beetham (L), Research Officer at the VCFG, Peter MacCallum Cancer Centre.
Dr Kevin Sek, loading the Incucyte SX5 Live Cell instrument at the Victorian Centre For Functional Genomics (VCFG) at Peter MacCallum Cancer Centre.
Human MCF7 adherent breast tumour cells where incubated alone or in the presence of CAR T cells in the Incucyte. Images were taken every 4 hours over 48 hours. MCF7 cells were stained red with NucLight and the number of live red stained tumour cells were counted over time. Cell death was measured through Caspase 3/7 green flourescence.

‘ Through our collaboration with Prescient Therapeutics, we wanted to determine the anti-tumour capabilities of Prescient’s new universal OmniCAR T cell platform. I teamed up with the VCFG staff who were critical in optimizing and setting up real-time, coculture experiments on the Incucyte. Using a small number of CAR T cells and tumour targets, I was able to obtain high quality videos and images, quantify tumour cell killing and T cell proliferation, and also determine multiple antigen targeting of the Universal OmniCAR T cells in a mixed tumour killing assay.”

Dr Kevin Sek
Dr Kevin Sek, Postdoctoral Researcher at the Peter MacCallum Cancer Centre.

The IncuCyte SX5, at the Victorian Centre for Functional Genomics (VCFG), is a live-cell imaging microscope that is housed inside a cell culture incubator. It allows researchers to image up to six 384-well plates in phase, green and red channels every 2 hours, in parallel! Its user friendly software can quantify cell number, size, shape and fluorescence intensity at the single-cell level over time. 

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