Caco-2 Permeability

The Caco-2 system has become the standard format to relate drug permeability to oral drug absorption. The Caco-2 cell model exhibits morphological as well as functional similarities to human intestinal enterocytes.

Features of Caco-2 cell monolayers:

  • Express many of the transporters found in the small intestine (bile acid, amino acid, dipeptide, tri-peptide, etc.)
  • Express the p-glycoprotein efflux pump and other ABC transporters
  • Exhibit metabolizing enzymes on the brush border membrane and in the cytosol
  • Form tight junctions between cells

Caco-2 permeability assays could be useful to predict

  • Transport mechanisms — passive diffusion
  • Transport mechanisms — carrier-mediated
  • Assessment of prodrugs
  • pH effects on transport

Assay Methodology

Quintara Discovery offers a validated Caco-2 permeability assay to assess transport in both directions, apical to basolateral (A-B) and basolateral to apical (B-A), across the cell monolayer. The bi-directional format of the assay allows to understand any involvement of a transport systems on the efflux of a test compound.

Caco-2 Permeability: Standard Assay Conditions (Customizable)

Testing Compound Concentration
10 μM

Assay Buffer
HBSS, pH 7.4 (or pH 6.5) on apical side; HBSS, pH 7.4 on basolateral side, 1-3% BSA optional on the receiving side

Plate Format
24-well or 96-well Transwell plates

Transport Period
1 hour at 37C

Detection Method
LC-MS/MS

Control Compounds
Digoxin

Deliverables
Papp value for A-B and B-A directions, efflux ratio

Turnaround Time
5-14 Days

Compound requirements
20 μL 10 mM DMSO stock