MTS viability: HeLa + doxorubicin (9-pt dose)

{
    "experiment": {
        "id": 314,
        "custom_id": "VIA-DOX-2026-0604",
        "title": "MTS viability: HeLa + doxorubicin (9-pt dose)",
        "template": "Cell Viability Assay",
        "template_version": "v3.2",
        "category": "Cell Biology",
        "status": "Complete",
        "started_on": "2026-11-04",
        "tags": [
            "cell-viability",
            "MTS-assay",
            "doxorubicin",
            "HeLa",
            "cytotoxicity"
        ]
    },
    "main_text": "This template captures an MTS-based cytotoxicity assay of doxorubicin against HeLa cervical carcinoma cells in a 96-well format, generating a 9-point dose-response curve and EC50 estimate.\nGoal: measure the cytotoxic effect of doxorubicin (anthracycline DNA-intercalator) on proliferating HeLa cells over a log-scale 9-point dose range (10 µM → 1.5 nM, 3-fold serial dilution), with 4 replicate wells per dose, and extract EC50, Hill slope, and assay window (Z’) for QC.\nMethod overview: 5,000 HeLa cells seeded per well in 100 µL DMEM + 10% FBS in a Corning 96-well flat-bottom plate, 24 hours pre-treatment. Doxorubicin diluted from 10 mM DMSO stock in growth media to 2× working concentrations, with vehicle control matched at 0.1% DMSO. 100 µL of 2× drug added to wells for 1× final concentration. 24 hours treatment at 37°C / 5% CO₂. 20 µL CellTiter 96 AQueous One Solution (MTS + PES) added per well, incubated 4 hours, absorbance read at 490 nm on plate reader. Blank-subtracted viability calculated relative to vehicle control wells. 4PL fit performed in GraphPad Prism.\nExpected: Sigmoidal dose-response with EC50 in the 0.3-0.6 µM range for HeLa (published range), Hill slope 1.0-1.8, R² > 0.99, Z’ > 0.5. Background < 0.1 A490; vehicle-control viability normalized to 100%; high-dose viability < 10%.",
    "extra_fields": {
        "Cells": {
            "Cell line": {
                "type": "select",
                "value": "HeLa, MCF7, A549, HCT116, primary culture"
            },
            "Seed density": {
                "type": "number",
                "value": "5000 cells/well"
            },
            "Pre-treatment time": {
                "type": "number",
                "value": "24 h"
            }
        },
        "Compound": {
            "Compound": {
                "type": "text",
                "value": "Doxorubicin"
            },
            "High dose": {
                "type": "number",
                "value": "10 µM"
            },
            "Dilution factor": {
                "type": "number",
                "value": "3"
            },
            "Dose points": {
                "type": "number",
                "value": "9"
            },
            "Replicates": {
                "type": "number",
                "value": "4"
            },
            "Treatment time": {
                "type": "number",
                "value": "24 h"
            }
        },
        "Readout & QC": {
            "Readout": {
                "type": "select",
                "value": "MTS, AlamarBlue, CellTiter-Glo, trypan blue"
            },
            "EC50": {
                "type": "number",
                "value": "0.41 µM"
            },
            "Z′-factor": {
                "type": "number",
                "value": "0.82"
            }
        }
    },
    "attached_files": [
        {
            "filename": "plate_layout.pdf",
            "type": "pdf",
            "size": "224 KB"
        },
        {
            "filename": "reader_A490_raw.csv",
            "type": "csv",
            "size": "18 KB"
        },
        {
            "filename": "dose_response_4PL.xlsx",
            "type": "xlsx",
            "size": "92 KB"
        }
    ],
    "steps": [
        {
            "n": 1,
            "description": "Trypsinize HeLa stock + count: 4.8 × 10⁵ cells/mL in DMEM+10%FBS",
            "duration": "0:10",
            "complete": true,
            "note": "95% viability by trypan blue"
        },
        {
            "n": 2,
            "description": "Seed 100 µL/well = 5,000 cells/well in inner 60 wells, outer rows H₂O",
            "duration": "0:15",
            "complete": true
        },
        {
            "n": 3,
            "description": "Incubate plate 24 h at 37°C / 5% CO₂",
            "duration": "24:00",
            "complete": true
        },
        {
            "n": 4,
            "description": "Prepare doxorubicin 9-point 3-fold dilution series in media (2× final)",
            "duration": "0:15",
            "complete": true,
            "note": "High 20 µM → low 3 nM (2×)"
        },
        {
            "n": 5,
            "description": "Add 100 µL of 2× drug to wells, vehicle-only to control wells (0.1% DMSO)",
            "duration": "0:10",
            "complete": true
        },
        {
            "n": 6,
            "description": "Incubate plate 24 h at 37°C / 5% CO₂",
            "duration": "24:00",
            "complete": true,
            "note": "Treatment 24 h"
        },
        {
            "n": 7,
            "description": "Add 20 µL CellTiter 96 AQueous One per well, swirl gently",
            "duration": "0:03",
            "complete": true
        },
        {
            "n": 8,
            "description": "Incubate 4 h at 37°C / 5% CO₂",
            "duration": "4:00",
            "complete": true,
            "note": "Color develops to orange-brown"
        },
        {
            "n": 9,
            "description": "Read A490 on plate reader; export CSV",
            "duration": "0:03",
            "complete": true,
            "note": "BioTek Synergy H1 · run 06:04:14:32"
        },
        {
            "n": 10,
            "description": "Calculate blank-subtracted viability + 4PL fit in Prism, extract EC50 + Z'",
            "duration": "0:25",
            "complete": true,
            "note": "EC50 0.41 µM · Z' 0.82 · R² 0.998"
        }
    ],
    "spreadsheet": [
        [
            "Dose (µM)",
            "Rep 1",
            "Rep 2",
            "Rep 3",
            "Rep 4",
            "Mean A₄₉₀",
            "% Viability"
        ],
        [
            "10",
            "0.121",
            "0.118",
            "0.115",
            "0.122",
            "0.119",
            "5"
        ],
        [
            "3.3",
            "0.140",
            "0.135",
            "0.142",
            "0.138",
            "0.139",
            "8"
        ],
        [
            "1.1",
            "0.226",
            "0.218",
            "0.232",
            "0.221",
            "0.224",
            "22"
        ],
        [
            "0.37",
            "0.508",
            "0.512",
            "0.495",
            "0.519",
            "0.509",
            "50"
        ],
        [
            "0.12",
            "0.788",
            "0.795",
            "0.781",
            "0.792",
            "0.789",
            "80"
        ],
        [
            "0.041",
            "0.872",
            "0.879",
            "0.865",
            "0.880",
            "0.874",
            "88"
        ],
        [
            "0.014",
            "0.925",
            "0.918",
            "0.931",
            "0.924",
            "0.924",
            "93"
        ],
        [
            "0.0045",
            "0.948",
            "0.939",
            "0.951",
            "0.945",
            "0.946",
            "95"
        ],
        [
            "0.0015",
            "0.962",
            "0.955",
            "0.968",
            "0.959",
            "0.961",
            "97"
        ],
        [
            "vehicle",
            "0.992",
            "1.005",
            "0.985",
            "1.000",
            "0.996",
            "100"
        ],
        [
            "blank",
            "0.072",
            "0.068",
            "0.074",
            "0.070",
            "0.071",
            "—"
        ]
    ],
    "links": {
        "experiments": [
            {
                "ref_id": "EXP-2026-0531",
                "title": "HeLa working stock + growth curve QC",
                "owner": "J. Chen",
                "date": "May 31"
            },
            {
                "ref_id": "EXP-2026-0608",
                "title": "Doxorubicin + cisplatin combo screen",
                "owner": "J. Chen",
                "date": "Jun 8"
            }
        ],
        "resources": [
            {
                "ref_id": "SOP-0184",
                "title": "MTS Cell Viability SOP v1.4",
                "owner": "QA Team",
                "date": "Jan 24"
            },
            {
                "ref_id": "KIT-0042",
                "title": "CellTiter 96 AQueous One Solution insert",
                "owner": "Promega Docs",
                "date": "Jun 2"
            },
            {
                "ref_id": "CPD-0341",
                "title": "Doxorubicin datasheet + Pubmed references",
                "owner": "Compound DB",
                "date": "Apr 8"
            }
        ]
    },
    "compounds": [
        {
            "name": "CellTiter 96 AQueous One Solution (MTS + PES)",
            "catalog_lot": "G3580 · Lot 0000465812",
            "stock_status": "In stock"
        },
        {
            "name": "Doxorubicin HCl (10 mM in DMSO)",
            "catalog_lot": "D1515 · Lot SLBR9421A",
            "stock_status": "In stock"
        },
        {
            "name": "DMSO (cell culture grade, vehicle ctrl)",
            "catalog_lot": "D2650 · Lot SLBT8847A",
            "stock_status": "In stock"
        },
        {
            "name": "DMEM + GlutaMAX (high glucose)",
            "catalog_lot": "10566016 · Lot 2722164",
            "stock_status": "In stock"
        },
        {
            "name": "FBS, heat-inactivated (HI-FBS)",
            "catalog_lot": "16140071 · Lot 2722084",
            "stock_status": "Low"
        }
    ],
    "storage": [
        {
            "name": "Doxorubicin stock + 10× working aliquots",
            "location": "-20 °C · Freezer C-2 · Drawer 1 · Box CPD-DOX-2026",
            "count": "×8"
        },
        {
            "name": "CellTiter 96 reagent (5 mL working)",
            "location": "-20 °C protected from light · Freezer C-2",
            "count": "×2"
        },
        {
            "name": "Plate reader raw data + Prism workbook",
            "location": "Cloud · /viability/2026-06/DOX-001/",
            "count": "×3"
        }
    ],
    "permissions": {
        "visibility": [
            {
                "type": "Team",
                "name": "Chen Lab"
            },
            {
                "type": "Person",
                "name": "J. Chen"
            }
        ],
        "can_write": [
            {
                "type": "Person",
                "name": "J. Chen"
            },
            {
                "type": "Person",
                "name": "Dr. M. Liu"
            }
        ]
    }
}