changes: use durvalumab as control
experiments:
Across All:
- Cell Counts consistent on input
- multi-concentration
- low medium high
Gains / Saturation (P0)
- secondary antibody titration → hypothesis: we are saturating our detectors and need to fine tune the secondary ab amount
- manual run for control
- suggest: add a fluorophore control - fluorescent beads to be a consistent marker
- suggest: manual run with same exact settings
Plate Washer (P0)
- Isolated experiment (plate washer → cytation) → isolates any plate washer issues from cytometer / cell count issues
- yellow dye
- reach out to Sothira
- use different plate washers (gg deck + 405s upstairs)
- no spin + wash run → addresses all hypotheses relating to plate washer variation
- just 1 wash
- plate flip for spin / wash → hypothesis: there’s some pin to pin variation down the columns of the plate washer (whether on aspirate or dispense pins)
- cytometer run (plate #2) gives related informaiton
Cytometer Settings (P1/P2) - defer for now - will test after Plate Washer tests
- time to read / order
- vary read order (spiral, groups of 3, reverse, etc - from Sothira’s tests)
- read same plate multiple times
- vary cycles before rinse, cycles before
- stop conditions
- cell counts + volume stop conditions
- gates, potentially as well
- cell counts + volume stop conditions
Cell Counts
Plan manual plate + secondary antibody titration (Bhavaani, Ajai, Noemi) outputs:
- gains for PE channel, V445 channel
- secondary antibody concentration for histogram centering Plate Washer isolated exp (Saahas, Andrew) outputs:
- uniformity on 056 plate washer with:
- 1x aspirate
- 1x dispense
- 1x aspirate + dispense
- 2x aspirate + dispense
- 4x aspirate + dispense
- 6x aspirate + dispense
- repeat above with different plate washers to test