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Fig. 2 | Stem Cell Research & Therapy

Fig. 2

From: Discrete adipose-derived stem cell subpopulations may display differential functionality after in vitro expansion despite convergence to a common phenotype distribution

Fig. 2

Flow cytometric and gating strategy using the Ex640 - Em664/22 nm channel. a Spherotech 6 peak Rainbow beads were used to identify a useful range of PMT potential. The bracket indicates voltage investigated during ensuing optimization. b PMT linearity and CV from the same experiment. c The narrowed range of PMT voltages was analyzed for optimal S/N using BD Biosciences Compensation beads conjugated with the labeled antibodies used in this study. The voltage that produced the highest S/N with no apparent double negativity was then chosen for each channel and its respective antibody (in box). *Double negativity. d To ensure the reproducibility of the experimental setup, the Spherotech single peak Rainbow beads were analyzed for 5000 events in each channel every time an experiment was conducted. e For sorting, the cells were first gated using SSCHeight vs. time plot to control for the stability of the flow cytometer. f The noise and debris were removed using FSCHeight vs. SSCHeight contour plot. g The doublets were discriminated and removed using FSCWidth vs. FSCHeight contour plot. h The live cells were identified by means of a viability stain. Abbreviations: PMT photomultiplier, CV coefficient of variation, S/N signal-to-noise ratio, AGM average geometric mean, GMFI geometric mean of fluorescence intensity, SSC side scatter, FSC forward scatter

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