A dissociated organoid is a suspension of single cells with different forward scatter (FSC) — roughly proportional to cell diameter — and different intensity on a fluorescence channel (FL1) from an antibody or reporter bound to a lineage marker.
FSC ∝ cell diameter (size)
FL1 ∝ [marker] × antibody/reporter brightness
Gate: keep event if FSC ≥ FSC_gate AND classify
positive if FL1 ≥ FL1_gate, else negative
Past the laser interrogation line, a real droplet cell sorter charges each droplet positive or negative according to the gate decision, then deflects it between two charged plates — exactly a CRT-style projectile in a uniform field, drawn here in 2D side-on cross-section:
while inside the plates (0 ≤ t ≤ t1):
y(t) = ½ (qE/m) t²
vy(t) = (qE/m) t
after the plates, no more field — straight line:
y(t) = y(t1) + vy(t1) (t − t1)
Here q/m is folded into the Deflection voltage slider (E = V/d across the plates), so raising the voltage bends the stream further into the top (positive) or bottom (negative) collection tube; uncharged waste events fall straight down the middle.
- Purity = (events in the positive gate that are truly the marker's target cell type) ÷ (all events in the positive gate).
- Yield = (target cells successfully captured in the positive gate) ÷ (all target cells that ever entered the stream).
- Switching the marker channel between LGR5-GFP and Villin-RFP flips which population lights up — the same physical gate can enrich for stem/progenitor cells or for differentiated cells depending on which reporter you are reading.
- Drag the scene to pan, scroll/pinch to zoom — the physics keeps running in world coordinates regardless of view.
Real-world relevance: this is the same FACS (fluorescence-activated cell sorting) workflow used to characterize and purify organoid-derived cell populations for the "Phenotyping and omics" step of organoid research — flow cytometry, paired with RNA-seq or proteomics, is how labs confirm what cell types an organoid actually grew.