CHIP PREPARATION
Organ-on-chip success begins with proper surface preparation. PDMS chips require plasma treatment or ECM coating (collagen I, fibronectin, or Matrigel) to enable cell attachment. Coat channels with 50-100 μg/mL ECM protein overnight at 4°C, then aspirate excess before cell seeding. Verify channel patency with media flush.
CELL SEEDING PROTOCOL
- Density: Typically 1-5 × 10⁶ cells/mL depending on channel dimensions
- Method: Inject cell suspension slowly to avoid air bubbles and shear damage
- Attachment: Invert chips or use static culture 2-4 hours for ceiling seeding
- Multi-layer: Seed endothelium first, allow attachment, then epithelium on membrane
- Verification: Image 24h post-seeding to confirm confluence and morphology
FLOW OPTIMIZATION
- Shear Stress: 0.1-1 dyn/cm² for endothelium, lower for epithelium
- Flow Rate: Calculate from channel dimensions and target shear stress
- Pulsatility: Peristaltic pumps provide physiological pulsatile flow
- Bubble Prevention: Degas media, use bubble traps, prime tubing completely
- Ramp-Up: Start static, increase flow gradually over 24-48h
ASSAY READOUTS
Common readouts include barrier function (TEER measurement, permeability), effluent analysis (albumin, urea, cytokines by ELISA), imaging (live/dead, immunofluorescence), and functional assays (CYP450 activity, transporter function). Establish baseline measurements before compound exposure for proper controls.