Healthy skin culture

Drug Discovery Assay – reference number: B117

This experiment assesses whether test articles cause inflammatory cytokine release in healthy skin, with lipopolysaccharide (LPS) as a reference compound. It uses healthy skin punch biopsies to explore test article effect on the expression on inflammatory cytokines via ELISA and/or rtPCR.

Testing Information

Introduction

The specific results that will be provided are on the inflammatory effect of test articles in human fresh skin.

Turnaround Time

For a standard experiment with n=3, tissue procurement is estimated to be 8-10 weeks. These timelines are a guide based on standard tissue criteria; any client-imposed alterations to the criteria may alter these estimates.

Test Article Requirements

Test article(s) to be provided by the Sponsor in storable aliquots at required test concentrations with information on diluent vehicle used. Stock solutions are prepared in deionized water unless otherwise requested. Sponsor to provide sufficient test article to run the entire study.

Suggested Testing

We suggest test conditions are assessed in triplet as a minimum.

Study Outline

Rationale and Experimental Design

Option 1: Systemic administration of test article

Full-thickness skin biopsies are achieved via punch-biopsy of residual surgical tissue. Biopsies are taken at a size of 3mm² and transferred into fortified media, leaving the epidermis exposed to air. Biopsies are incubated in optimum conditions for a maximum of 72 hrs. Tissue media is collected for phenotype analysis by ELISA and/or the biopsy set is collected for rtPCR.

An example of the conditions assessed for 3 test articles are detailed below (it is recommended a minimum of triplicates should be used for each condition):

• Control
• Test article 1
• Test article 2
• Test article 3
• Positive control

Option 2: Topical application of test article

Full-thickness skin biopsies are achieved via punch-biopsy of residual surgical tissue. Biopsies are taken at a size of 8mm² and transferred into specially fortified media, leaving the epidermis exposed to air. Biopsies are incubated in optimum conditions for a maximum of 72 hrs. Tissue media is collected for phenotype analysis by ELISA and/or the biopsy set is collected for rtPCR.

An example of the conditions assessed for 3 test articles are detailed below (it is recommended a minimum of triplicates should be used for each condition):

• Control
• Test article 1
• Test article 2
• Test article 3
• Positive control

Exclusion Criteria

No specific exclusion criteria are in place other than to reject macroscopically diseased/necrotic tissue.

Standardization and Qualification

Tissue that does not respond to stimulation with the positive control would be deemed non-functional.

Analysis

Cytokine analysis: Supernatant samples can be analyzed for specific analytes of your choice by a multiplex ELISA platform. Each analyte will be quantified by interpolation against a standard curve generated on the same 96 well analysis plate.

Other forms of end point analysis are available such as gene expression or immunohistochemistry.

Example data from Healthy skin culture model

Normal Skin Histopathology Maintained for up to One Week in Culture

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