Skip to content
← Back to browse
LIVER Publication-derived

Human Liver Organoid Screening Platform for DILI Risk Prediction

Source Zhang et al., 2023 · University of Michigan · 10.1016/j.jhep.2023.01.019

👤 Charles J. Zhang, Sophia R. Meyer, Matthew J. O'Meara, Sha Huang, Meghan M. Capeling, Daysha Ferrer-Torres, Charlie J. Childs, Jason R. Spence, Robert J. Fontana, Jonathan Z. Sexton ⏱ 14 days 📋 10 phases 🧫 Human iPSC (fibroblast-derived)

Abstract

This protocol describes the generation and characterization of human liver organoids (HLOs) derived from iPSC lines in two complementary formats: dispersed HLOs in 384-well plates for high-throughput screening and intact HLOs adapted to a liver-on-chip system (PaDLOC) for enhanced physiological assessment. Both platforms enable identification of hepatotoxic compounds and prediction of synergistic drug toxicities through biochemical assays, morphological profiling, and single-cell transcriptomics.

Cell source
Human iPSC (fibroblast-derived)
Application
Disease modeling and drug screening for drug-induced liver injury (DILI) risk prediction

Protocol overview

35 steps across 10 phases

Preparation of PaDLOC (Patient-Derived Liver-On-Chip) System Pre-seeding
  1. 1 Prepare Chip S1 channels with extracellular matrix
  2. 2 Prepare dispersed HLO cell suspension
PaDLOC Cell Seeding and Attachment Day 0
  1. 1 Seed cells into bottom channel
  2. 2 Allow bottom channel cell attachment
  3. 3 Seed cells into top channel
  4. 4 Allow top channel cell attachment
  5. 5 Wash both channels with hepatocyte growth media
PaDLOC Culture with Flow Days 1–7
  1. 1 Attach chip to Pod Portable Module
  2. 2 Place loaded pods into Zoë Culture Module
  3. 3 Introduce media flow at constant rate
  4. 4 Maintain media and culture for 7 days pre-treatment
PaDLOC Drug Treatment Days 8–14
  1. 1 Replace media with drug-containing media
  2. 2 Maintain constant flow during 7-day treatment period
  3. 3 Collect media outflow at designated time points
  4. 4 Measure ALT, AST, and albumin in collected media
PaDLOC Imaging and Morphological Analysis Day 14 (post-treatment)
  1. 1 Fix and stain PaDLOC cells
  2. 2 Acquire confocal microscopy images
  3. 3 Analyze morphological features
PaDLOC Single-Cell RNA Sequencing Day 14 (post-treatment)
  1. 1 Harvest cells from treated PaDLOCs
  2. 2 Perform single-cell RNA sequencing
  3. 3 Analyze differential expression
384-Well Dispersed HLO Culture and Drug Screening Days 0–5
  1. 1 Coat 384-well plates with collagen type I
  2. 2 Seed dispersed HLO cells into 384-well plates
  3. 3 Allow cells to adhere
384-Well Drug Dispensing and Dose-Response Assessment Days 2–5
  1. 1 Prepare compound stocks
  2. 2 Dispense compounds using digital dispenser
  3. 3 Incubate cells with compounds
384-Well Fixation and Staining Day 5 (post-treatment)
  1. 1 Fix cells in 384-well plates
  2. 2 Permeabilize and block
  3. 3 Stain with fluorescent dyes and antibodies
  4. 4 Wash and prepare for imaging
384-Well Automated Confocal Imaging and Analysis Day 5–6 (post-staining)
  1. 1 Acquire confocal microscopy images
  2. 2 Perform image segmentation and feature extraction
  3. 3 Quantify cell viability and calculate IC50 values
  4. 4 Perform dimensionality reduction and clustering

Full SOP

🔬

Create a free account to access this protocol

Join OrganMatch to unlock step-by-step procedures, reagent concentrations, QC checklists, and downloadable batch record templates.

Create free account

Already registered? Log in

Attribution

This SOP was authored by Organthis based on the published method in Zhang et al., 2023. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

This wording is awaiting legal review.

Something wrong with this entry? Report an issue with this protocol

Need a commercial licence?
Use this protocol in your therapeutic or diagnostic pipeline.
Request Commercial Licence