Platform Technologies

At REPROCELL, we offer a unique range of experimental techniques for translating data from preclinical species to humans. These specialized instruments have been used by our human tissue scientists for over two decades, and by pharmacologists for even longer. If you want to find out more about the laboratory equipment used in our drug discovery studies, we have included more information about these systems below.

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Tissue  baths

  • Muscle contractility 
  • Nerve-muscle interaction 
  • Vaso-dilation/ constriction
  • Broncho-dilation/ constriction

Ussing chambers

  • Drug transport, metabolism, and absorption
  • Ion channel function

Ex vivo tissue culture

  • Cytokine and chemokine release 
  • Inflammation  

3D tissue models

  • Marker expression
  • Cell migration 
  • Drug transport, metabolism, and absorption

Ex vivo tissue culture

Our scientists offer a range of fresh ex vivo tissue culture (organoculture) systems that can be customized to suit your requirements. By using small tissue explants that reflect human biology, we can retain phenotypic relevance and maximize throughput per donor. Throughput per donor varies depending on the tissue type but can be as high as 20-24 for skin explants or higher still for precision-cut lung slices (PCLS). Biospecimens are cultured for up to seven days, allowing a range of functional endpoints to be measured, such as cytokine or chemokine production, gene expression, and immunohistochemistry.

These organotypic assays can use healthy or diseased biospecimens ethically sourced from patients with psoriasis, inflammatory bowel disease, and COPD. When combined with the analysis of up to 50 analytes per well, testing your compounds in human fresh tissues provides powerful data in a cost-effective manner. REPROCELL-04.06.18_0304

Endpoint analysis 

By combining multiplex immunoassays with our ethical access to human fresh healthy and diseased tissue samples, REPROCELL offers a unique service based on the Bio-Plex Immunoassay system, which is capable of reading up to 50 analytes per sample. 

The measurement of analytes such as growth factors, chemokines, or cytokines released from tissue samples can be a powerful pharmacodynamic experiment, particularly in diseased biospecimins. Our Luminex platform provides a sensitive measurement of a wide range of analytes and is compatible with multiplex kits from several manufactures including Bio-Rad, Life Technologies, and Millipore. REPROCELL-04.06.18_0296-2

Precision Cut Lung Slice (PCLS) Assays

PCLS assays retain the structural and functional integrity of lung tissue to offer a phenotypically accurate model for lung behavior. This model can be used to measure a variety of endpoints such as structural integrity, tissue viability, or biomarker release. By comparing animal and human data, the translation of preclinical animal data into human clinical information can be assessed. Typical workflow for a PCLS project. Other human tissues can also be prepared as slices.

Diagram showing the set up for precision-cut lung slices or PCLS

Diagram showing the setup of our PCLS assays.

Skin Disease Models

Fresh human skin is cultured in media and a psoriasis-like phenotype is induced using a proprietary cocktail of compounds and test compounds are applied in order to test their ability to reduce inflammation. After stimulation of the Th17 pathway, the inhibition of inflammation by potential new drugs is assessed by determining the release of cytokines, such as IL-17 or IL-22, using a Luminex assay.

The set up for a typical full-thickbness human skin tissue explant experiment-1

Diagram showing the setup of our ex vivo skin culture assays. 


Ussing Chambers

Ussing chambers are a flexible, cost-effective means of investigating mucosal drug transport or drug-mediated effects on tissue behavior. This system is used for investigations into the electrochemical properties of membranes and epithelia, most commonly the mucosae of the gastrointestinal tract or respiratory system. The setup consists of two fluid-filled, heated, gassed chambers, which are separated only by the mucosal tissue preparation. Typically, 6 to 12 isolated tissue preparations are used per donor, allowing multiple drugs to be investigated in each experiment.

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Diagram showing the setup of our Ussing chamber assay

Endpoint analysis 

The Ussing chamber system allows investigations in intact mucosal tissues including the electrical, metabolic, and absorptive properties of the membrane. Endpoints measured include in vitro permeability of drug compounds (Papp), phase I and II metabolism (e.g. CYP3A4 mediated metabolism of drugs), short-circuit current (Isc), potential difference (PD) measurements, and transepithelial resistance (TEER).

Ion channel function

When measuring ion channel function, it is common to record, in real-time, the potential difference (V), transepithelial resistance (Ω/cm2), and short-circuit current (mAs). Changes in ion channels (e.g. CFTR) or ion exchange mechanisms (e.g. Na-K-Cl cotransporters) result in a change in the short-circuit current. This is of major interest in mucosal biospecimins because of the role of ion channels in regulating fluid secretion. REPROCELL-04.06.18_0243-2

Drug absorption, transport, and metabolism

With respect to drug absorption, transport and metabolism, Ussing chambers are recognized as the gold standard. By allowing investigations in whole tissues a phenotypically accurate balance of drug-metabolizing enzymes and drug transporters can be maintained.

To measure the passage of the drug and any metabolites, a typical experiment involves the application of a drug to the apical surface of the tissue, followed by periodic sampling of the apical and basolateral chambers. Oral bioavailability is dependent on a number of factors, two key factors being gastrointestinal permeability and metabolism. 

A review by Astra Zeneca highlighted the benefits of human fresh tissue mounted in Ussing chambers for the accurate prediction of oral bioavailability. Alternative methods such as Caco-2 cells or intestinal microsomes are useful for either permeability or metabolism experiments, but not both. Moreover, using human fresh gastrointestinal tissue avoids any species differences in oral bioavailability. REPROCELL-04.06.18_0236

Drug-mediated changes or release of mediators

This type of experimental readout once again involves the addition of test compounds to the apical and/or basolateral surfaces of the tissue. However, in this case, periodic samples collected from the bathing fluid and processing of the tissue offer a flexible method to investigate drug-mediated changes in tissue biology.

An increasingly common application of Ussing chambers is to measure the release of gastrointestinal peptides such as glucagon-like-peptide-1 (GLP-1): an incretin released from intestinal epithelial cells, in response to nutrients, that stimulates insulin release from the pancreas. REPROCELL-04.06.18_0246-1

Organ baths and wire myographs

Organ baths and wire myographs are versatile in vitro pharmacology techniques that are a mainstay of pharmacology. Many important discoveries have been made using these systems, including the experiments used to discover endothelium-derived relaxing factor (EDRF), executed by Nobel prize winner Professor Robert Furchgott.

REPROCELL offers the most advanced instrumentation on which to conduct organ bath and wire myograph experiments. Many tissue types can be used to measure drug behavior, including blood vessels, airways, gastrointestinal tissues, genitourinary tissues, and cardiac muscle.

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Diagram showing the setup of our wire myograph system 

Organ Baths

Organ baths have been used for decades by pharmacologists to investigate nerve-muscle interactions and the contractility of smooth, cardiac, or skeletal muscles. In this system, isolated tissues are suspended within a fluid-filled, heated, and gassed chamber. Changes in tissue contractility are recorded via sensitive force transducers. Typical endpoints measured using this system include vasoconstriction, bronchodilatation, gastrointestinal motility, or bladder activity. Organ baths are suitable for a wide range of tissue types and sizes, from strips of muscle down to small airways or blood vessels ~2mm diameter.

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Diagram showing the setup of our organ bath system 

Respiratory and cardiac contractility 

At REPROCELL, we have numerous Panlab organ baths at our laboratories in Glasgow and Maryland, with experienced scientists on hand to investigate the contractility of multiple isolated tissues at a time.

For example, multiple bronchi may be isolated from lung tissue to allow comparisons between the broncho-relaxant properties of compounds to treat asthma or COPD. The propensity of test compounds to cause vasoconstriction of coronary arteries, cardiac muscle contractility, or the nerve-mediated control of gastrointestinal tissues can be studied in organ baths. REPROCELL-04.06.18_0192

Wire Myographs

Wire myographs are smaller versions of organ baths that allow investigation of the contractile and relaxant properties of isolated blood vessels and airways. They were originally developed by Professor Mike Mulvaney to sensitively measure vasoactivity of resistance arteries, however, they can also be used to investigate small airways. Tissues are mounted in the myograph horizontally between two stainless steel 'jaws' held in place by thin wires. As the tissue contracts or relaxes the change in force is transmitted via one of the wires to a sensitive isometric force transducer, capable of detecting micro-Newton forces.

This means that very small arteries, such as those vital to the control of organ-specific blood flow and the overall vascular component of blood pressure, can be studied ex vivo. Months of practice are required to dissect tissues without damage, and a high level of operator skill is vital.  REPROCELL-04.06.18_0163

3D tissue models using Alvetex™

For over 15 years REPROCELL’s CSO, Professor Stefan Przyborski, has harnessed our award-winning Alvetex 3D scaffold technology to test pharmaceutical compounds in skin, liver, and neuronal tissues. By combining the human relevance of our iPSC-derived cell lines with Alvetex technology, our team can develop a solution for your 3D in vitro assay needs.

Because human cells exist in three dimensions (3D) in vivo, culturing them in a two-dimensional (2D) format results in loss of form and function. 3D tissue models have repeatedly demonstrated a more accurate response to drugs, chemicals, and cosmetics than their 2D counterparts because they:

  • More closely mimic human tissue micro-anatomy
  • Are more phenotypically accurate e.g. maintenance of CYP450 activity in 3D in comparison with 2D culture
  • Allow you to interrogate the mechanisms of action of your compound with confidence
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Making an Inquiry

Whatever tissue or species you are working on, REPROCELL can help translate your findings to humans. View our catalog of assays in human isolated functional tissues, or contact our scientists today to discuss a customized approach that will demonstrate the efficacy, safety, and ADME properties of your compounds.

What happens when I make an inquiry? 

  • We will send you a fully-costed proposal and timeline.
  • You wend your test compounds to REPROCELL.
  • REPROCELL will perform the assay in accordance with agreed protocols and on completion of the work will supply raw data and a written report detailing results.
  • Tissues and test compounds can be returned to you for further analysis.

 

REPROCELL will participate at

BioPharm America 2021 conference

20 – 23 Sept, 2021 / Virtual Meeting