# ECL Command Center · Sickbay Padd Protocol Outline
# STAMP: ecl_sickbay_protocol_outline.txt · 2026-09-15 · py=0
# STATUS: OUTLINE ONLY · full SLL requires ECL account + sample ship · not executed
# REF: cro_methods_sheet.md · pilot_plate_map.tsv · JANE_MONDAY_GRAIN_ROADMAP_NOW

IF X = ...
  ECL = Emerald Cloud Lab · Command → Run → Explore → Analyze
  SLL = Symbolic Lab Language (Wolfram-based) · typed in Command Center notebook

================================================================================
PHASE 0 · COMMAND (before any Run)
================================================================================
1. Register project: "Sickbay-TNBC-PacR-Pilot" · PI Brian King · ORCID on file
2. Ship cell lines to ECL Austin (15500 Wells Port Drive) · defined lots L1/L2/L3
3. Import pilot_plate_map.tsv as Container/plate model in Command Center
4. Link each Sample[L1|L2|L3] to container wells per TSV

================================================================================
PHASE 1 · RUN — Look 1 (WALK · plate reader)
================================================================================
ECL instrument classes (typical):
  · Automated cell culture + 96-well seeding
  · Liquid handler: paclitaxel dilution series 0.1–1000 nM
  · Incubator 72 h · 37 C · 5% CO2
  · Plate reader: CellTiter-Glo / CellTox Green

SLL-style outline (pseudocode · not validated on ECL):
  Experiment["Look1_Walk_Safety"];
  Samples = {L1, L2, L3};
  Plate = Import["pilot_plate_map.tsv"];
  DoseSeries = Range[0.1 nM, 1000 nM, LogSpace];
  For each well in Plate[look==1]:
    Transfer[Sample[well.line], well, Density -> 5000 cells/well];
    AddCompound[Paclitaxel, well.paclitaxel_nM];
    If[well.condition=="MbCD", AddCompound[MbetaCD, 1.2 mM]];
  Incubate[72 Hour];
  ReadLuminescence[Plate]; ReadFluorescence[Plate, CellToxGreen];
  Gate["IC50_MCF10A >= 5*IC50_231PacR"] -> If fail, Abort["HALT"];

================================================================================
PHASE 2 · RUN — Look 2 (RIVER · efflux)
================================================================================
ECL instrument classes:
  · Fluorescence plate reader · kinetic read
  · Rhodamine-123 load 30 min · efflux chase

  Experiment["Look2_Efflux"];
  Require[Look1.Pass];
  Wells = Plate[look==2];
  LoadDye[Rhodamine123, 5 uM, 30 Minute];
  ReadKinetic[efflux, Interval -> 5 Minute, Duration -> 60 Minute];
  Arm MbCD: AddCompound[MbetaCD, 1.2 mM] at t=0;
  Gate["J_after/J_before < 0.5"];

================================================================================
PHASE 3 · RUN — Look 3 (PORCH · confocal)
================================================================================
ECL instrument classes:
  · Confocal / high-content imaging
  · Fixed t* = 1.0 h post-MβCD

  Experiment["Look3_Cluster"];
  Require[Look2.Pass];
  Stain[Pgp_ABCB1, Alexa488];
  Stain[Raft, CTB_AF488];
  Fix[t_star -> 1 Hour post MbCD];
  Image[ZStack, 63x];
  Analyze[ClusterRatio, threshold -> C/N < 0.10];

================================================================================
PHASE 4 · RUN — Look 4 (SINK · LC-MS/MS)
================================================================================
ECL instrument classes:
  · Compound treatment · wash · scrape/lyse
  · Mass spectrometry (LC-MS/MS) · quant paclitaxel ng/mg protein

  Experiment["Look4_Retention"];
  Require[Look3.Pass];
  Treat[Paclitaxel, 100 nM, 1 Hour];
  Wash[3x PBS];
  Quench[ice cold methanol];
  Extract[protein assay + LC-MS/MS];
  Gate["retention >= 2x vehicle in L1"];

================================================================================
PHASE 5 · EXPLORE + ANALYZE (Constellation)
================================================================================
  · Attach all raw files to knowledge graph nodes (well → sample → instrument → run)
  · Export gate table CSV + SHA-256 manifest
  · Optional: share read-only link with collaborator (not public patient data)

================================================================================
LIMITS (honest)
================================================================================
  · This outline is NOT submitted to ECL · no SLL compile test on Forge
  · gotokell CRO links may 500 until redeployed · source of truth = hitme/kids/docs/
  · OpenBind spores = inventory labels · not part of this wet protocol
  · Helpers not doctors · no clinical claim

CONTACT ECL: business team demo via emeraldcloudlab.com/get-started
WORD=amen · ecl · sickbay · outline · protocols