BD FACSMelody™ Cell Sorter Easy Experiment Setup

Start a new experiment by selecting the fluorochromes to be used. For this two-color bead sort, FITC and PE are selected, though the BD FACSMelody can accommodate up to nine different fluorochromes. Optionally add marker names such as CD4.

Set up four initial graphs for sample analysis and add additional plots including a 50/50 dot plot and contour plot. Load the sample onto the instrument and use pink sliders to adjust instrument settings until the bead population becomes visible.

Use pink sliders to fine-tune voltage settings on the FITC and PE graphs. The BD FACSMelody software automatically recalculates compensation, which comes pre-calculated for up to 11 different fluorochromes.

Record 1000 events from the pre-sort sample. Then create gates around the PE-positive and FITC-positive populations, labeling them as P1 (PE positive) and P2 (FITC positive) for downstream sorting.

Select 5 mL tubes as collection vessels and add buffer volume. Set the sort parameters to collect 25,000 beads per tube, directing FITC-positive beads to the left tube and PE-positive beads to the right tube.

Position collection tubes and click Start Sort. The instrument calculates the estimated sort time based on flow rate and cell concentration; increase speed to reduce the ~30 minute sorting duration in real time.

Once sorting completes, the system confirms the collected event count. Name the sorted sample, review the sort report showing efficiency and collection statistics, and export the report as a PDF for record keeping.

Unload the collection tube from the instrument. Select either daily or long-term shutdown mode to properly power down the BD FACSMelody cell sorter.