EA202 – Bidirectional Caco-2

>>EA202 – Bidirectional Caco-2

EA202 – Bidirectional Caco-2

$673.60

EA202 – ExpressPlus Bidirectional Permeability Through Caco-2 Cell Monolayers

This assay is used to determine the permeability of a test article through Caco-2 cell
monolayers in the apical-to-basolateral and basolateral-to-apical direction

Catalog number EA202

Required from Sponsor

  • Either a minimum of 300 µL of test article at 10 mM in DMSO, or 5 mg of powder
  • Molecular mass (exact mass) of the test article and its salt form
  • MSDS or handling and storage information, e.g., light sensitive, store at -20°C, etc.

Deliverables

  • Cell batch QC results
  • The percent recovery of the test article from the assay wells containing Caco-2 cell monolayers
  • The apparent permeability (Papp) in both directions
  • The absorption potential of a test article classified as either low or high using the following criteria for the apical-to-basolateral (A→B) direction:
    • Low: Papp <1.0 x 10-6 cm/s
    • High: Papp ≥1.0 x 10-6 cm/s
  • The efflux ratio (Papp B→A)/(Papp A→B)
    • The efflux is classified as significant when: Efflux ≥2.0 and Papp (B→A) ≥ 1.0 x 10-6 cm/s

Substrate

  • Test article at 5 µM in HBSSg with maximum DMSO concentration ≤ 0.8%

Assay System

  • Confluent monolayers of Caco-2 (clone C2BBe1) cells, 21 to 28 days old

Assay Conditions

  • Bidirectional permeability of the test article in Caco-2 cells
  • Transport buffer: HBSSg, pH 7.4 ± 0.2
  • Receiver well containing 1% BSA
  • Apical and basolateral side at pH 7.4
  • Dose two cell monolayers (n=2)
    • Dose apical side for (A→B) assessment
    • Dose basolateral side for (B→A) assessment
  • Sample from both apical and basolateral sides at 120 minutes
  • Determine the concentrations of test article using a generic LC-MS/MS method with a minimum 6-point calibration curve

Assay QC

  • The quality of the monolayer batch is verified using control compounds before the monolayers are released for use
  • The quality of each monolayer used in the assay is verified by calculating the Papp for the control article, lucifer yellow, dosed post-experimentally

Notes
1. The results from this assay are sent to the sponsor in the ExpressPlus report format, which may include graphical representations of data and comparison with historical data for reference articles.
2. For reproducible results the solubility of the test article must be greater than 5 µM in HBSSg with 1% DMSO.