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Transwell based BBB Model Service

Introduction Transwell-based BBB Model Workflow What We Can Offer Case Study FAQ Related Sections
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Introduction

The Blood-Brain Barrier (BBB) is a key hurdle in neurological drug development, with 98% of small molecules failing to penetrate the CNS. Creative Biolabs' Transwell-based BBB Model uses a co-culture of brain endothelial cells and astrocytes, a functional barrier with validated tight junctions, enabling precise drug permeability and mechanistic studies.

As a validated, high-fidelity platform, it provides human-relevant permeability and transport data to de-risk CNS candidates, offering actionable solutions to development challenges and accelerating compounds through preclinical stages cost-effectively.

Transwell-Based BBB Model

The Transwell based BBB Model, particularly when enhanced by Creative Biolabs' proprietary co-culture protocols, is a robust platform for both preclinical screening and mechanistic discovery.

Model Construction

Cell Selection: Endothelial cells with high tight junctions (e.g., hCMEC/D3, bEnd.3) are cultured in the Transwell upper compartment to simulate the BBB endothelial barrier. Astrocytes (e.g., U251) are seeded in the lower compartment to mimic in vivo BBB characteristics, supporting barrier tightness and functionality.

Culture Conditions: The Transwell membrane is pre-coated with rat tail collagen and fibronectin for cell adhesion. Endothelial cells are cultured in appropriate medium (e.g., 1640 medium with 10% FBS and 1% PS) at 37°C, 5% CO₂. Medium is changed daily for 6-7 days to form a tight monolayer.

A schematic diagram of the time for inoculating bEnd.3 and C8-D1A cells on the apical and lateral basal sides of Transwell to obtain a contact co-culture in vitro BBB model. (OA Literature)Fig.1 bEnd.3 and C8-D1A cells were inoculated on the apical and lateral basal sides of Transwell to obtain a contact co-culture in vitro BBB model.1

Applicable Disease Models

The established BBB model serves as a foundation for studying the onset and progression of various neurological disorders where BBB dysfunction is a key factor. Applicable models include:

  • Neurodegenerative Conditions: Alzheimer's Disease (AD) and Parkinson's Disease (PD), particularly for investigating how peripheral factors (like erythrocyte-derived extracellular vesicles) interact with and compromise the neurovascular unit.
  • Acute Injuries: Traumatic Brain Injury (TBI) and Ischemic Stroke, for assessing compounds designed to protect or restore barrier integrity following acute insults.
  • Inflammatory Disorders: Modeling systemic inflammation to study subsequent BBB breakdown and immune cell extravasation into the CNS.

Advantages

  • Simplicity and Cost-Effectiveness: Format is more accessible/scalable than microfluidic devices, enabling cost-effective high-throughput screening. Accelerates CNS lead identification by cutting the cost per data point.
  • Quantitative TEER Measurement: Provides direct, non-invasive electrical measurement of barrier integrity for validation/QC. Offers a quantitative metric to confirm barrier "tightness" in permeability assays.
  • Astrocyte End-Feet Mimicry: Porous membrane enables endothelial-astrocyte contact, promoting physiological tight junctions. Replicates cell-cell communication for superior barrier function/integrity.

Workflow

Our streamlined process ensures transparency, high-quality results, minimized project turnaround time, and maximized data integrity. This service typically takes 4-7 weeks, depending on compound structure complexity and the total number of test articles.

Required Starting Materials

  1. Test Compound Specifications: Detailed structure, purity data, and required working concentrations for the assay.
  2. Solubility and Stability Data: Information regarding the compound's stability in the assay media and solvent compatibility.
  3. Target Transport Mechanisms: Any preliminary data or hypothesis regarding the compound's transport (e.g., passive diffusion vs. carrier-mediated transport).
Step Activities Involved
Initial Consultation & Design Scope definition, material transfer agreement, and assay parameter setting (e.g., time points, apical/basolateral dosing).
Cell Culture & Barrier Establishment Co-culture of brain endothelial cells (bEnd.3 or iPSC-derived) and astrocytes on the Transwell membrane.
Permeability Assays Dosing the compound in the apical (blood side) and basolateral (brain side) chambers; collecting samples at predetermined time intervals.
Efflux Pump Characterization Using specific P-gp inhibitors (e.g., verapamil) as controls to assess the role of active efflux in compound transport.
Data Analysis & Reporting Advanced calculation of transport kinetics, data visualization, and correlation with industry standards.

Final Deliverables

  1. Comprehensive Permeability Report: A detailed document containing all raw data, calculated Papp values for both directional flows, and statistical analysis.
  2. Efflux Ratio and Liability Analysis: A clear determination of the Efflux Ratio for each compound, categorizing the risk associated with active efflux.
  3. Tight Junction Immunostaining Data: High-resolution microscopy images and data validating the maintenance of tight junction proteins (ZO-1, Claudin-5) throughout the experiment.

What We Can Offer

Creative Biolabs understands that your drug discovery pipeline requires flexibility and uncompromising data quality. Our Transwell based BBB Model service is built to be a true partner in your CNS R&D, providing validated, scalable solutions at every stage.

One-Stop Translational Service

Seamless support from small-scale feasibility studies and pilot screens (Tier 1) up to large-scale High-Throughput Screening (HTS) campaigns (96-well format) for lead identification and optimization.

Guaranteed Barrier Fidelity

We operate under a robust Quality System, with mandatory validation steps including consistent, high Transendothelial Electrical Resistance (TEER) measurement and verification of key tight junction protein expression (e.g., ZO-1, Claudin-5) before any compound testing.

Process Analytic Techniques (PAT) for Stability

Continuous monitoring and strict aseptic procedures throughout the culture process ensure maximum cell viability and stable, predictable barrier function over the full duration of your assay.

Customized Modeling and Cell Sourcing

We offer true customization, including flexible assay run modes (e.g., single-pass vs. cumulative sampling), and the integration of your specific iPSC-derived cells or disease-specific patient models for unparalleled translational relevance.

Comprehensive Efflux & Uptake Analysis

Beyond standard Papp screens, we provide detailed mechanistic assays to quantify the involvement of multiple CNS transporters (P-gp, BCRP, etc.), giving you the full transport profile of your compound.

Detailed, Audit-Ready Documentation

Every project concludes with a final report that meets high industry standards, ensuring the data is reliable, traceable, and suitable for internal review and regulatory submissions.

Experience the Creative Biolabs Advantage - Get a Quote Today

Case Study

Transendothelial Electrical Resistance Analysis FITC Dextran Permeability Assay
TEER analysis was performed on the resistance of Transwell membranes, where bEnd.3 and C8-D1A cells were grown or co-cultured using chopstick electrodes. (OA Literature)
Fig.2 TEER analysis was performed using chopstick electrodes to record the resistance on the Transwell membrane.1 		The barrier ability of the in vitro BBB model was analyzed by the isothiocyanate fluorescein glucan permeability assay. (OA Literature)
Fig.3 The barrier ability of the in vitro BBB model was analyzed by the FITC dextran permeability assay.1
Solvent Persistence Solvent Leakage Tests
Compared with single-cell culture models, the co-cultured Transwell has the highest solvent persistence. (OA Literature)
Fig.4 Co-cultured Transwell has the highest solvent persistence.1 		Compared with single-cell culture models, the co-cultured Transwell has the lowest solvent leakage value. (OA Literature)
Fig.5 The solvent leakage test was used to measure the time for which bEnd.3, C8-D1A and co-cultured cells could retain the culture medium at the top of the Transwell.1

Customer Reviews

[Superior P-gp Validation]

Using Creative Biolabs' Transwell-based BBB Model in our lead optimization research significantly improved the accuracy of efflux liability predictions. The confirmed P-gp functional data aligned perfectly with follow-up in vivo rodent models, saving over six months of unnecessary synthetic chemistry work.

Ana R. Carter
[High TEER Consistency]

The ability to achieve consistent, high TEER values with Creative Biolabs' Transwell-based BBB Model facilitated our study of small molecular transport across the barrier. This tight barrier lets us distinguish subtle permeability differences among analogs that previous leakier systems failed to detect.

David S. Mitchell
[Translational Relevance]

The option to use iPSC-derived cells via Creative Biolabs' Transwell-based BBB Model was key to our investigation into neurodegenerative pathways. The model accurately reproduced tight junction (ZO-1 and Claudin-5) compromise under specific inflammatory stimuli, providing highly translational mechanistic data.

Joan G. Reynolds

FAQs

A: Monocultures often yield non-physiologically high permeability (Papp) values due to a lack of cell-to-cell signaling and low TEER. Our co-culture model achieves a significantly tighter barrier, confirmed by high TEER and functional P-gp, resulting in data that is far more predictive of in vivo transport kinetics.
A: Absolutely. While P-gp is a core capability, our models can be adapted to assess the involvement of other key transporters (e.g., BCRP, MRPs) through the use of specific inhibitors and relevant cell lines. We encourage you to discuss your specific transporter targets in your inquiry so we can customize the assay.
A: A standard, non-toxicology-focused permeability assessment for up to 10 compounds typically requires 4 to 5 weeks from receipt of the test articles to final report delivery. Larger projects or those requiring complex mechanistic endpoints will be slightly longer.
A: We prioritize translational relevance. While we have rigorously validated iPSC-derived lines available, we are happy to discuss incorporating or validating your specific iPSC lines for endothelial and/or astrocyte differentiation to create a model perfectly tailored to your patient cohort research.
A: Every batch of cells is subject to strict quality control, including phenotypical characterization and, crucially, a mandatory barrier function check. We only proceed to client dosing once the barrier has achieved a minimum TEER threshold and passed positive control permeability and efflux assays.

The Creative Biolabs Transwell based BBB Model represents a critical advancement in preclinical neurological research, offering an unparalleled balance of high-throughput scalability and physiological fidelity. By accurately modeling the human neurovascular unit, including functional P-gp and co-culture synergy, we provide the predictive data necessary to confidently advance or terminate your CNS drug candidates.

Contact Our Team for More Information and to Discuss Your Project

Reference

  1. Park, Jun Sung, et al. "Establishing co-culture blood–brain barrier models for different neurodegeneration conditions to understand its effect on BBB integrity." International Journal of Molecular Sciences 24.6 (2023): 5283. https://doi.org/10.3390/ijms24065283. Distributed under Open Access license CC BY 4.0, figures were cropped.

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