Providing single-cell sensitivity, real-time flow cytometry is uniquely suited to quantify multimodal receptor functions of live cultures. Using adult neural progenitor cells, the P2X7 receptor function was assessed via calcium influx detected by calcium indicator dye, transmembrane pore formation by ethidium bromide uptake, and phagocytosis using fluorescent latex beads.
Total time
~4–6 hours per experiment (cell preparation, staining, flow cytometry acquisition, and analysis)
Model organism
Adult neural progenitor cells (mammalian)
Steps
1
Prepare single-cell suspension of neural progenitor cells
Dissociate and prepare adult neural progenitor cells into a single-cell suspension optimized for flow cytometry analysis, ensuring cell viability and appropriate concentration for downstream measurements.
▶ 00:58
2
Measure calcium influx by live-cell flow cytometry
Load live cells with calcium indicator dye and use real-time flow cytometry to detect and quantify P2X7-mediated calcium influx in response to receptor activation.
▶ 02:11
3
Measure pore formation by live-cell flow cytometry
Apply ethidium bromide uptake assay to detect transmembrane pore formation induced by P2X7 receptor activation, using flow cytometry to quantify fluorescent signal in single cells.
▶ 05:21
4
Measure phagocytosis by live-cell flow cytometry
Incubate cells with fluorescently labeled latex beads and use flow cytometry to quantify bead uptake as a readout of P2X7-dependent phagocytic activity.
▶ 07:02
5
Analyze multimodal P2X7 receptor function data
Integrate flow cytometry data from calcium influx, pore formation, and phagocytosis measurements to characterize comprehensive P2X7 receptor signaling in neural progenitor cells.
▶ 09:02