Presentation description

This project investigates the efficiency of HeLa cells in taking up HIV-1 plasmids when transfected using standard Lipofectamine transfection protocols. To measure the number of plasmids, a stochastic counting algorithm was used, transfecting a varying ratio of Dark, Cherry, and GFP non-infectious HIV backbones into cells while keeping the total DNA concentration constant. Fluorescence-activated cell sorting (FACS) on a CytoFLEX S flow cytometer was employed to measure the fluorescence of individual cells in both GFP and Cherry channels in a sample of 100,000 cells. The CytoFLEX S, equipped with multiple lasers and detectors, allowed for precise detection and quantification of the fluorescent signals emitted by the transfected cells. We programmed a simulation of the CytoFLEX S data to analyze and simulate different Cherry and GFP dilutions. Our results show that individual HeLa cells take up between 10 and 300 backbones, with this distribution best represented by three Gaussians with the following means and standard deviations: (25, 2), (100, 3), and (300, 4). We also identified additional sources of noise contributing to our measurements and compared our values to similar distributions measured in 293 cells using a different technique.