Assays

MB Research works closely with pharmaceutical companies and government agencies around the world to help ensure the safety and efficacy of new medicines being brought to market. MB Research provides an extensive array of testing protocols that will help meet your regulatory requirements to ensure safe use of your products.

OECD Assays

In Vitro Assays

EpiOcular™ Eye Irritation Test (EIT)
OECD 492, GHS
The purpose of this study is to provide classification of chemicals concerning their eye irritation potential using an alternative to the Draize Rabbit Eye Test, according to the OECD Test Guideline No. 492, “Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Identifying Chemicals Not Requiring Classification and Labelling for Eye Irritation or Serious Eye Damage”. The EpiOcular™ EIT is intended to differentiate those materials that are UN GHS No Category (i.e., do not meet the requirements for UN GHS classification) from those that would require labeling as either UN GHS Category 1 or 2.
EpiDerm™ Skin Corrosion Test (SCT)
OECD 431, GHS
The purpose of this study is to provide classification of the dermal corrosion potential of chemicals by using a three-dimensional human epidermis model, according to the OECD Guideline for the Testing of Chemicals No. 431, “In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method”. The EpiDerm™ SCT allows discrimination between non-corrosive and corrosive substances in accordance with U.N. GHS classification. Corrosivity classification can be further divided into either optional Sub-category 1A or a combination of Sub-categories 1B and 1C.
Corrositex®
OECD 435, GHS

The Corrositex® assay is performed using a kit produced and distributed by In Vitro International. The kit contains tubes of proprietary buffers, a Chemical Detection System (CDS), and components used to make synthetic proteinaceous macromolecular bio‐barriers. After preliminary testing to determine if the test article is compatible with the Corrositex® assay (qualification) and to determine cut‐off times (categorization), the test article is applied topically on prepared bio‐barriers set atop vials of CDS. The amount of time it takes for the test article to penetrate the bio‐barriers and cause a visually detectable change in the CDS (breakthrough time) can be used to determine the UN Packing Group or the UN GHS Subcategory (classification).

EpiDerm™ Skin Irritation Test (SIT)
OECD 439, GHS
The purpose of this study is to provide classification of the dermal irritation potential of chemicals by using a three-dimensional human epidermis model, according to the OECD Guideline for the Testing of Chemicals No. 439, “In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method”. The EpiDerm™ SIT allows discrimination between irritants (Category 2) and non-irritants, in accordance with U.N. GHS classification.
3T3 Neutral Red Uptake Phototoxicity Test
OECD 432

The assay is designed to detect the phototoxicity induced by the combined action of a test article and light by using an in vitro cytotoxicity assay with the Balb/c 3T3 mouse fibroblast cell line. The test identifies aqueous-soluble compounds (or formulations) that have the potential to exhibit in vivo phototoxicity.

KeratinoSens™
OECD 442D

Upon exposure to skin sensitizers, the KeratinoSens™ Test measures activation of Keap1-Nrf2-antioxidant/electrophile response element (ARE). These tests use an immortalized, adherent, human keratinocyte cell line (HaCaT) that was transfected with a plasmid to monitor luciferase gene induction.

Human Cell Line Activation test (h-CLAT)
OECD 442E, GHS
To determine the skin sensitization potential of a test article due to cell activation by the measurement of changes in the expression of dendritic cell surface markers (CD86, CD54) via flow cytometry. The THP-1 human monocytic cell line serves as the test system. The h-CLAT is recommended by EURL ECVAM as part of an Integrated Approach to Testing and Assessment (IATA) to support the discrimination between sensitizers (i.e., UN GHS Category 1) and non-sensitizers for the purpose of hazard classification and labeling, per OECD Guideline 442E “In Vitro Skin Sensitization: Human Cell Line Activation Test (h-CLAT).”
Direct Peptide Reactivity Assay (DPRA)
OECD TG 442C

The DPRA is an in chemico method that quantifies cysteine‐ or lysine‐containing peptide depletion following 24 hours incubation with the test article (TA). Relative peptide concentration is measured by HPLC with gradient elution and UV detection at 220 nm. Cysteine and lysine peptide percent depletion values are then calculated and used in a prediction model, which allows assigning the TA to one of four reactivity classes used to support the discrimination between sensitizers and sonosensitizers.

Alternative Assays

Bovine Corneal Opacity & Permeability (BCOP) Assay
OECD 437, GHS

Freshly harvested bovine corneas from the eyes of cattle are utilized in the BCOP test method in order to evaluate the ocular irritancy and corrosive potential of a test chemical by assessing quantitative measurements of: 

(1) Changes in corneal opacity, measured with an opacimeter

(2) Corneal permeability, measured by a visible light spectrophotometer

Both measurements are used to calculate an In Vitro Irritation Score (IVIS), which is used to assign a classification category.

Bovine Corneal Opacity & Permeability Test and EpiOcular™ Eye Irritation Test Combination Study (BCOP-EIT)
OECD 437, OECD 492, GHS

The objective of this study is to determine the potential for ocular irritation using OECD Guidelines for the Testing of Chemicals, Nos. 492 and 437, which serve as alternatives to the traditional Draize methodology. The use of two internationally-validated, OECD-accepted non-animal tests can provide a GHS categorization for the vast majority of test articles. The combination of the Bovine Corneal Opacity and Permeability Test and EpiOcular™ Eye Irritation Test followed by a Weight-of-Evidence analysis of the results allows for the classification of GHS Category 2.

Bacterial Reverse Mutation Test (Ames Test)
OECD 471
The bacterial reverse mutation test evaluates the mutagenic potential of a test article by using Salmonella typhimurium and Escherichia coli strains to detect point mutations, which includes the substitution, addition or deletion of one or a few DNA base pairs, in the presence and absence of metabolic activation (S9).
Local Lymph Node Assay (LLNA) Flow Cytometry
OECD 429
To determine the sensitizing potential of topically applied test material utilizing the LLNA, and measurement of lymphocyte proliferation by BrdU incorporation (by flow cytometry). Our protocol is not a guideline assay but is designed to be compliant with OECD TG 429.
Local Lymph Node Assay (LLNA-BrdU ELISA)
OECD 442
The local Lymph Node Assay (LLNA) is an alternative study to the Guinea Pig Sensitization Test, used for determining sensitization potential of materials. Following exposure to a sensitizing test substance, lymphocyte proliferation occurs in the local lymph node. The LLNA measures increased proliferation of lymphocytes in the auricular lymph nodes (which drain the site of exposure; ears). Proliferation is assessed by determining the incorporation of the thymidine analog, bromodeoxyuridine (BrdU), into the DNA of lymph node cells using flow cytometric method.

Additional Assays

Acute Oral Toxicity – Up and Down Procedure (UPD)
OECD 425
The purpose of this study is to determine the potential for toxicity of the test article when administered orally. This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guidelines, OCSPP 870.1100, and in the Organisation for Economic Co-operation and Development (OECD) Guidelines for the Testing of Chemicals, Guideline 425.
Acute Oral Toxic Class Determination
OECD 423
The study is designed to determine the potential for toxicity of a test article when administered orally. This method uses pre-defined doses and the results allow the test article to be classified according to the Globally Harmonized System for the classification of chemicals which cause acute toxicity.
Acute Dermal Toxicity / LD50
OECD 402

The purpose of this study is to determine the potential of toxicity of the test article when applied dermally. This study is designed to comply with the standards set forth of in the current Organisation for Economic Co-operation and Development (OECD) Guideline for the Testing of Chemicals, No. 402.

Acute Dermal Irritation and Corrosion
OECD 404
The study is designed to determine the skin irritancy or corrosive potential of a test article. This determination is made by treating the skin of up to three test subjects for a three-minute, one-hour and/or four-hour exposure period and evaluating the skin at scheduled time points for up to 14 days.
Acute Inhalation Toxicity LC50 of Aerosols
OECD 403
This study assesses the acute inhalation toxicity of a respirable, aerosolized test article following a four-hour exposure.
Delayed Contact Dermal Sensitization Test – Buehler Method
OECD 406
Contact dermal sensitization is an immunological process where the host animal, through repeated skin exposure, acquires a specific allergic sensitivity to a substance. In the Buehler model, contact dermal sensitivity is manifested as increased erythema.
Guinea Pig Maximization Test (GPMT)
OECD 406
Contact dermal sensitization is an immunological process where the host, through repeated skin exposure, acquires a specific allergic sensitivity to a substance. In the Guinea Pig Maximization (GPMT) model, contact dermal sensitivity is manifested as increased erythema.

EPA-OCSPP Assays

Oral Toxicity

Acute Oral Toxicity – Up and Down Procedure (UPD)
OCSPP 870.1100

The purpose of this study is to determine the potential for toxicity of the test article when administered orally.  This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guidelines, OCSPP 870.1100, and in the Organisation for Economic Co-operation and Development (OECD) Guidelines for the Testing of Chemicals, Guideline 425.

Dermal Toxicity

Acute Dermal Toxicity / LD50
OCSPP 870.1200

The purpose of this study is to determine the potential for toxicity of the test article when applied dermally for a 24-hour exposure. This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guideline OCSPP 870.1200: Acute Dermal Toxicity and U.S. Department of Transportation (DOT) 49 CFR 173.132(b)(2).

Inhalation Toxicity

Acute Inhalation Toxicity
OCSPP 870.1300

The purpose of this study is to provide information on health effects that may arise from short term exposure by the inhalation route. This study is designed to comply with the standards set forth in U.S. Environmental Protection Agency (EPA) Health Effects Test Guideline, OCSPP 870.1300 and The Organisation for Economic Co-operation and Development (OECD) Guideline for the Testing of Chemicals No. 403.

Ocular Irritation

Acute Ocular Irritation and Corrosion
OCSPP 870.2400

This study assesses the potential of a test article to cause irritation or corrosion to the eye by evaluation for up to 21 days following a single exposure. The intent of this study is to evaluate test articles that have been identified as not severely irritating or corrosive to the eye according to the OECD-recommended method: the Bovine Corneal Opacity and Permeability Assay (BCOP).

Dermal Irritation

Acute Dermal Irritation and Corrosion
OCSPP 870.2500

The study is designed to determine the skin irritancy or corrosive potential of a test article.  This determination is made by treating the skin of up to three test subjects for a three-minute, one-hour and/or four-hour exposure period and evaluating the skin at scheduled time points for up to 14 days. 

Dermal Sensitization

Delayed Contact Dermal Sensitization Test – Buehler Method
OCSPP 870.2600

The purpose of this study is to determine the potential of a product to promote skin sensitization reactions after repeated applications using the method of Ritz and Buehler, 1980, Current Concepts in Cutaneous Toxicity. This study is designed to comply with the standards set forth in U.S. Environmental Protection Agency (EPA) Health Effects Test Guideline OCSPP 870.2600: Skin Sensitization, and The Organisation for Economic Co-operation and Development (OECD) Guideline for the Testing of Chemicals No. 406: Skin Sensitization.

Guinea Pig Maximization Test (GPMT) Magnusson-Kligman
OCSPP 870.2600

Contact dermal sensitization is an immunological process where the host animal, through repeated skin exposure, acquires a specific allergic sensitivity to a substance. In the Guinea Pig Maximization (GPMT) model, contact dermal sensitivity is manifested as increased erythema.

Local Lymph Node Assay (LLNA-BrdU ELISA)
OCSPP 870.2600

The purpose of this study is to determine the sensitizing potential of topically applied test articles. This LLNA protocol, utilizing the BrdU ELISA method, is designed to be a stand-alone assay for dermal sensitization as described in the NIH report “The Murine Local Lymph Node Assay: A Test Method for Assessing the Allergic Contact Dermatitis Potential of Chemicals/Compounds,” NIH No. 99-4494, and the LLNA test guidelines as defined in EPA OCSPP 870.2600 and OECD Guideline for the Testing of Chemicals No. 442B.

ISO-USP Assays

In Vitro Assays

Test by Direct Contact (MEM Elution)
ISO 10993-5 & ISO 10993-12, USP 23
This assay tests for biocompatibility. The test article is extracted with DMEM media supplemented with 5% fetal bovine serum and tested on monolayer L929 mouse fibroblast cells based on the current ISO 10993‐5 standard. This protocol is a qualitative test to determine the potential of a test article to produce cytotoxicity.
Test by Indirect Contact (Agar Overlay)
ISO 10993-5
This assay tests for cytotoxicity by indirect contact. This assay is not appropriate for leachables that cannot diffuse through the agar or may react with agar. The test article is laid on an agar layer poured above a monolayer of L929 mouse fibroblast cells based on the current ISO 10993-5 standard. This protocol is a qualitative test to determine the potential of a test article to produce cytotoxicity.
Neutral Red Uptake (NRU) Cytotoxicity Test
ISO 10993-5
This assay tests for the cytotoxic potential of a test article when applied to a monolayer of epidermal keratinocytes or fibroblasts in culture.
KeratinoSens™
ISO 10993-10

Upon exposure to skin sensitizers, the KeratinoSens™ Test measures activation of Keap1-Nrf2-antioxidant/electrophile response element (ARE). These tests use an immortalized, adherent, human keratinocyte cell line (HaCaT) that was transfected with a plasmid to monitor luciferase gene induction.

Human Cell Line Activation Test (h-CLAT)
ISO 10993-10
The h‐CLAT is an in vitro method that quantifies changes in cell surface markers on a human cell line following 24 hours exposure to test chemical, as measured by flow cytometry. The relative fluorescence intensity of CD54+ and CD86+ expression when compared to solvent control are calculated and used in a prediction model to support the discrimination between sensitizers and non‐sensitizers.
In Vitro Reconstructed Human Epidermis Model (SIT)
ISO 10993-23
The SIT utilizes three-dimensional reconstructed human epidermis (RhE) tissues. The RhE tissues are composed of primary human cells, which have been cultured to form a stratified, highly differentiated squamous epithelium morphologically similar to that of human skin. Following exposure to a test article (TA) and incubation, tissue viability is measured via MTT reduction. The SIT assay will classify a liquid, powder or solid as either non-irritating or irritating. This assay is not designed to test gases and aerosols.

Alternative Assays

Bacterial Reverse Mutation Test (Ames Test)
ISO 10993-3
The bacterial reverse mutation test evaluates the mutagenic potential of a test article by using Salmonella typhimurium and Escherichia coli strains to detect point mutations, which includes the substitution, addition or deletion of one or a few DNA base pairs, in the presence and absence of metabolic activation (S9).
Local Lymph Node Assay (LLNA-BrdU ELISA)
ISO 10993-10
The local Lymph Node Assay (LLNA) is an alternative study to the Guinea Pig Sensitization Test, used for determining sensitization potential of materials. Following exposure to a sensitizing test substance, lymphocyte proliferation occurs in the local lymph node. The LLNA measures increased proliferation of lymphocytes in the auricular lymph nodes (which drain the site of exposure; ears). Proliferation is assessed by determining the incorporation of the thymidine analog, bromodeoxyuridine (BrdU), into the DNA of lymph node cells using flow cytometric method.

Additional Assays

Implantation in Subcutaneous Tissue
ISO 10993-6

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Implantation in Muscle
ISO 10993-6

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Guinea Pig Maximization Test (GPMT)
ISO 10993-10
The Guinea Pig Maximization Test (GPMT) uses a combined intradermal and topical induction regime to assess a test article’s ability to dermally sensitize the test subjects.  A challenge of the highest non-irritating concentration is used to elicit potential effects.
Closed-patch Test (Buehler Test)
ISO 10993-10
The Buehler Assay uses a three– or nine-induction regime to assess a test article’s ability to dermally sensitize the test subjects. A challenge of the highest non-irritating concentration is used to elicit potential effects.
Acute Systemic Toxicity
ISO 10993-11

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Pyrogenicity Test in Rabbits
ISO 10993-11, USP 151

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Irritation Test by Skin Exposure
ISO 10993-23

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Irritation Test by Intracutaneous (Intradermal) Administration
ISO 10993-23

For more information about our 10993-23 Assays, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

Special Irritation Tests
ISO 10993-23

For more information about our 10993-23 Special Irritation Tests, please contact our Client Services Team at clientservices@mbresearch.com or call 215-536-4110.

DOT Assays

Acute Dermal Irritation and Corrosion
DOT 49 CFR 173.137
The study is designed to determine the skin irritancy or corrosive potential of a test article. This determination is made by treating the skin of up to three test subjects for a three-minute, one-hour and/or four-hour exposure period and evaluating the skin at scheduled time points for up to 14 days.
Acute Dermal Toxicity / LD50
DOT 49 CFR 173.132 (a)(1)(ii), DOT 49 CFR 173.133(a)(1)
The purpose of this study is to determine the potential for toxicity of the test article when applied dermally for a 24-hour exposure. This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guideline OCSPP 870.1200: Acute Dermal Toxicity and U.S. Department of Transportation (DOT) 49 CFR 173.132(b)(2).

Data analysis will be performed in accordance with EPA Health Effects Test Guideline OCSPP 870.1000: Acute Toxicity Testing and DOT 49 CFR 173.132 (a)(1)(ii) and 49 CFR 173.133(a)(1).
Acute Oral Toxicity – Up and Down Procedure (UPD)
DOT 49 CFR 173.132(a)(1)(i)
The purpose of this study is to determine the potential for toxicity of the test article when administered orally. This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guidelines, OCSPP 870.1100, and in the Organisation for Economic Co-operation and Development (OECD) Guidelines for the Testing of Chemicals, Guideline 425.

Data analysis will be performed in accordance with the current EPA Health Effects Test Guidelines, OCSPP 870.1100, the Consumer Product Safety Commission (CPSC) issued pursuant to and for the implementation of the Federal Hazardous Substances Act, the current U.S. Department of Transportation (DOT), 16 CFR Part 1500.3(c)(2)(i)(A), 49 CFR 173.132(a), and/or in OECD Guidelines for the Testing of Chemicals, Guideline 423 and 425. Guideline 423 is referred to in OCSPP 870.1000 as an acceptable method to assess lethality within a dose range.

CPSC/FHSA Assays

Acute Dermal Irritation and Corrosion
FHSA 16 CFR 1500.41

The study is designed to determine the skin irritancy or corrosive potential of a test article.  This determination is made by treating the skin of up to three test subjects for a three-minute, one-hour and/or four-hour exposure period and evaluating the skin at scheduled time points for up to 14 days. 

Acute Inhalation Toxicity of Aerosols
FHSA 16 CFR 1500.3(c)(2)(i)(B)
The purpose of this study is to provide information on health effects that may arise from short term exposure by the inhalation route. This study is designed to comply with the Consumer Product Safety Commission (CPSC) standards regulated under the Federal Hazardous Substances Act (FHSA) set forth in 16 CFR 1500.3(c)(2)(i)(B).
Acute Oral Toxicity – Up and Down Procedure (UPD)
FHSA 16 CFR 1500.3(c)(2)(i)
The purpose of this study is to determine the potential for toxicity of the test article when administered orally. This study is designed to comply with the standards set forth in the current U.S. Environmental Protection Agency (EPA) Health Effects Test Guidelines, OCSPP 870.1100, and in the Organisation for Economic Co-operation and Development (OECD) Guidelines for the Testing of Chemicals, Guideline 425.

Data analysis will be performed in accordance with the current EPA Health Effects Test Guidelines, OCSPP 870.1100, the Consumer Product Safety Commission (CPSC) issued pursuant to and for the implementation of the Federal Hazardous Substances Act, the current U.S. Department of Transportation (DOT), 16 CFR Part 1500.3(c)(2)(i)(A), 49 CFR 173.132(a), and/or in OECD Guidelines for the Testing of Chemicals, Guideline 423 and 425. Guideline 423 is referred to in OCSPP 870.1000 as an acceptable method to assess lethality within a dose range.

OCSPP Genetic Toxicity Assays

Bacterial Reverse Mutation Test (Ames Test)
OCSPP 870.5100
The bacterial reverse mutation test evaluates the mutagenic potential of a test article by using Salmonella typhimurium and Escherichia coli strains to detect point mutations, which includes the substitution, addition or deletion of one or a few DNA base pairs, in the presence and absence of metabolic activation (S9).

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