Evaluation of the GARD assay in a blind Cosmetics Europe study

ALTEX Online first published February 17, 2017 https://doi.org/10.14573/altex.1701121

Johansson H., Gradin R., Forreryd A., Agemark M., Zeller K., Johansson A., Larne O., van Vliet E.,  Borrebaeck C., Lindstedt M.,


Chemical hypersensitivity is an immunological response towards foreign substances, commonly referred to as sensitizers, which gives rise primarily to the clinical symptoms known as allergic contact dermatitis. For the purpose of mitigating risks associated with consumer products, chemicals are screened for sensitizing effects. Historically, such predictive screenings have been performed using animal models. However, due to industrial and regulatory demand, animal models for the purpose of sensitization assessment are being replaced by animalfree testing methods, a global trend that is spreading across industries and market segments. To meet this demand, the Genomic Allergen Rapid Detection (GARD) assay was developed. GARD is a novel, cell-based assay that utilizes the innate recognition of xenobiotic substances by dendritic cells, as measured by a multivariate readout of genomic biomarkers. Following cellular stimulation, chemicals are classified as sensitizers or non-sensitizers based on induced transcriptional profiles. Recently, a number of animal-free methods were comparatively evaluated by Cosmetic Europe, using a coherent and blinded test panel of reference chemicals with human and local lymph node assay data, comprising a wide range of sensitizers and non-sensitizers. In this paper, the outcome of the GARD assay is presented. It was demonstrated that GARD is a highly functional assay with a predictive performance of 83% in this Cosmetics Europe dataset. The average accumulated predictive accuracy of GARD across independent datasets was 86%, for skin sensitization hazard. Keywords: GARD, sensitization, in vitro, predictive accuracy, alternative methods

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Testing Human Skin and Respiratory Sensitizers—What Is Good Enough?

Int. J. Mol. Sci. 2017, 18(2), 241; doi:10.3390/ijms18020241

Malmborg A., Borrebaeck C. A.K.


Alternative methods for accurate in vitro assessment of skin and respiratory sensitizers are urgently needed. Sensitization is a complex biological process that cannot be evaluated accurately using single events or biomarkers, since the information content is too restricted in these measurements. On the contrary, if the tremendous information content harbored in DNA/mRNA could be mined, most complex biological processes could be elucidated. Genomic technologies available today, including transcriptional profiling and next generation sequencing, have the power to decipher sensitization, when used in the right context. Thus, a genomic test platform has been developed, denoted the Genomic Allergen Rapid Detection (GARD) assay. Due to the high informational content of the GARD test, accurate predictions of both the skin and respiratory sensitizing capacity of chemicals, have been demonstrated. Based on a matured dendritic cell line, acting as a human-like reporter system, information about potency has also been acquired. Consequently, multiparametric diagnostic technologies are disruptive test principles that can change the way in which the next generation of alternative methods are designed.

genomics; skin sensitization; adverse outcome pathways; next generation in vitro tests

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SenzaGen and Bioglan working together for the next generation of test methods

SenzaGen has signed a cooperation agreement with the pharmaceutical company Bioglan, which develops and manufactures creams and ointments for the pharmaceutical and cosmetics industries. The collaboration means that Bioglan will test GARDskin for skin allergens in a number of creams that are included in various end products.

Bioglan is a pharmaceutical company that focuses on research, development and manufacture of medicines, especially semi-solids such as creams and ointments. Bioglan manufactures topical medicines and personal care products for customers within the pharmaceutical industry. The company works with everything from product development and clinical trials through to full-scale production. Safety testing and allergy testing are part of Bioglan’s everyday routine and today human skin patch tests are the preferred method of testing end products for skin allergies. SenzaGen’s GARDskin offers an alternative that may eventually replace these tests and therefore the two companies have now chosen to enter into a partnership where the use of GARDskin on complex end products will be further developed.

“With Bioglan we’ll benefit from an extensive exchange of knowledge and will have the opportunity to test GARDskin on several different formulations. We will also gain a deeper understanding of the extraction process, and can thus develop our products and processes further. Our goal is that GARDskin will work even on end products, and that more companies will therefore take note of our technology and realize that there are safer and better tests than the ones which are still routinely used in many places today,” says Anki Malmborg Hager, CEO of SenzaGen.

Bioglan, with its production in Malmö, seeks to depart from traditional testing methods on human skin and instead use the next generation’s GARD test for safety testing of the company’s products. GARDskin investigates and finds allergens in chemicals and pharmaceutical preparations through a unique genome technology that completely replaces the need for animal testing. The goal is that GARDskin should replace current tests, thus providing a better assessment of allergenicity and a more cost-effective approach.


For more information
Anki Malmborg Hager, CEO, SenzaGen AB                                 Birgitta Svensson, Head of Development
Email: amh@senzagen.com                                                            Email: birgitta.svensson@bioglan.se
Phone: +46 768 284822                                                                   Phone: +46 40-287561, +46 708-149354


About Bioglan

Bioglan AB is your partner from product development through to full-scale manufacturing. We offer a total solution for all your production and service needs. Bioglan welcomes both small and large challenges, and is your partner of choice for

  • One-off studies during product development
  • Production of material for clinical trials (phase I – III)
  • Commercial scale production for international markets

With more than twenty five years of experience in research, development and manufacturing of medicines, we can offer you all the services you need within the areas of semi-solids and liquids.

Next generation skin sensitisation testing.

Personal Care Magazine Europe, sept 2016

Malmborg Hager A., Johansson H., Lindstedt M., Borrebaeck CA.


DNA is the software that predicts human phenotypes. Genomics utilises DNA as such a piece of software, which in several recent applications has been demonstrated to be able to predict a number of biological features, enabling anything from face recognition to vaccine development. The tremendous information content, harboured in the DNA, should be harnessed in tests of cosmetic ingredients and formulations, to get a complete insight into what is happening in the body when these types of products are applied to the human skin. This type of complex information gives a holistic view of a human condition that many of us are familiar with, such as e.g. allergic contact dermatitis, and has the potential not only to classify chemicals used in cosmetics but also to determine the magnitude to which a
chemical affects the human body (the potency).

This distinguishes genomics from most other test principles, where in many cases only one or two markers are being
monitored, such as DC activation markers (CD86, CD54), genes involved in cytoprotective responses to oxidative stress or electrophilic compounds (Nrf2, Keap1), or proinflammatory cytokines, (e.g. IL18).

Next generation skin sensitisation testing. PC Magazine, Sept 2016. Malmborg Hager A., Johansson H., Lindstedt M., Borrebaeck CA.

From genome-wide arrays to tailor-made biomarker readout – Progress towards routine analysis of skin sensitizing chemicals with GARD.

Toxicol In Vitro. 2016 Dec;37:178-188. doi: 10.1016/j.tiv.2016.09.013. Epub 2016 Sep 13.

Forreryd A., Zeller K., Lindberg T., Johansson H., Lindstedt M


Allergic contact dermatitis (ACD) initiated by chemical sensitizers is an important public health concern. To prevent ACD, it is important to identify chemical allergens to limit the use of such compounds in various products. EU legislations, as well as increased mechanistic knowledge of skin sensitization have promoted development of non-animal based approaches for hazard classification of chemicals. GARD is an in vitro testing strategy based on measurements of a genomic biomarker signature. However, current GARD protocols are optimized for identification of predictive biomarker signatures, and not suitable for standardized screening. This study describes improvements to GARD to progress from biomarker discovery into a reliable and cost-effective assay for routine testing. Gene expression measurements were transferred to NanoString nCounter platform, normalization strategy was adjusted to fit serial arrival of testing substances, and a novel strategy to correct batch variations was presented. When challenging GARD with 29 compounds, sensitivity, specificity and accuracy could be estimated to 94%, 83% and 90%, respectively. In conclusion, we present a GARD workflow with improved sample capacity, retained predictive performance, and in a format adapted to standardized screening. We propose that GARD is ready to be considered as part of an integrated testing strategy for skin sensitization.


GARD; In vitro assay; Predictive genomic biomarker signature; Skin sensitization

Link to articel on line

Next generation skin sensitisation testing

Malmborg Hager A., Johansson H., Lindstedt M., Borrebaeck CA. Next generation skin sensitisation testing. PC Magazine, Sept 2016

GARD approved for validation within the OECD

GARD, Senzagen´s in vitro assay for sensitization, has been included in the OECD Test Guideline Programme (TGP) work plan with the TGP No  4.106. The decision was made at the WNT National Coordinators Test guideline programme meeting in the end of April. The validation process for GARD will now be initiated.

SenzaGen’s test Genomic Allergen Rapid Detection (GARD) has been reviewed by all OECD WNT (The Working Group of the National Coordinators of the Test Guidelines Programme (WNT)) task force members and has been included in the Test Guideline Programme. This means that the process of formally validating GARD can begin. The purpose of the validation process is to show that GARD is a reproducible assay that can be use by external laboratories. The assay will be transferred externally to perform a so called “ring trial”.

“This is a very important decision for the development of GARD as an international sensitization test and we are very confident we will have a successful ring trial. Even if our customers have been convinced by the quality of our test, it will make a big difference from a regulatory point of view when the test is validated”, says Anki Malmborg Hager.

The GARD test has been scientifically validated and has gone through successful in house validations prior to this with industrial partners. The test assay is based on a new innovative genomic technique which is argued to be the future of sensitization testing now that animal bans are more and more prevailing. As the chemical industry needs to find accurate and reliable tests, GARD will be an important player on the testing market once it has finalised its validation.

“The OECD WNT representatives were very interested and positive to the GARD test and to include it in the TGP work plan. The representatives also look forward to receiving information regarding the formal validation, when these results are available. The demand is high for new alternative test methods and it has been an interesting journey as a representative for Swedish Chemicals Agency (Kemikalieinspektionen) to present GARD to OECD as it has such great potential. I am very much looking forward to follow the future for this test”, says Henrik Appelgren, national coordinator at Kemikalieinspektionen.

Two external laboratories will now test GARD on several different chemicals to confirm that the test is reproducible. This process has started and final validation is expected within one year.


For more information:
Anki Malmborg Hager, CEO, SenzaGen AB
E-mail: amh@senzagen.com
Phone: +46 768 284822

Brian Rogers, CEO, SenzaGen Inc
E-mail: brian.rogers@senzagen.com
Phone: +1 530 304 7648


About GARD
By analyzing 200 and 389 markers, depending on the test, GARD generates massive amounts of data and delivers results with 90 % prediction accuracy. This can be compared to the golden standard, animal tests on mice, that provides 72 % prediction. SenzaGen’s test also has the ability to measure potency (strength) of a substance and can thus determine the degree to which a substance is an allergen.


About SenzaGen
SenzaGen provides dermal and respiratory in vitro testing for the cosmetic, chemical and pharmaceutical industries replacing the need for animal testing. The company’s unique test GARD is based on research from the Department of Immune Technology at Lund University. SenzaGen is based at Medicon Village in Lund, Sweden.

SenzaGen signs new customer – entry into the pharmaceutical industry

SenzaGen has signed a strategic contract with a major international pharmaceutical company. This means that SenzaGen’s test GARD will be used for the testing of drug candidates in early development. The agreement opens up a whole new market for GARD, which previously has been used primarily for testing in the cosmetics industry.

Based at Medicon Village in Lund, SenzaGen develops and implements animal-free allergy tests on chemicals and proteins. SenzaGen’s test GARD (Genomic Allergen Rapid Detection) has been developed over 15 years and measures significantly more parameters than existing tests on the market. This makes GARD a safer and more effective test than other available options.

So far, GARD has primarily been used for testing in the cosmetics industry to detect potential allergens in products, but it is now entering the pharmaceutical industry.

“Since the start, collaborations with pharmaceutical companies has been a major objective for SenzaGen. We see great potential in this market and we are looking forward to learning more about the industry, with an ambition to be able to develop and offer GARD to even more customers in the pharmaceutical industry”, says Anki Malmborg Hager, CEO SenzaGen

The main advantage of using GARD is that the test is based on genomic technology and is performed in vitro. This is the first time that a pharmaceutical customer uses SenzaGen’s method. The tests can be done in a very early phase of drug development, which helps pharmaceutical companies to develop safer drugs at faster pace and at a lower cost.


For more information:
Anki Malmborg Hager, CEO, SenzaGen AB
E-mail: amh@senzagen.com
Phone: +46 768 284822


About SenzaGen
SenzaGen provides in vitro testing for the cosmetic, chemical and pharmaceutical industries replacing the need for animal testing. The company’s unique test GARD is based on research from the Department of Immune Technology at Lund University. SenzaGen is based at Medicon Village in Lund, Sweden and has 10 employees.

About GARD
By analyzing 200 and 389 markers, depending on the test, instead of 1-2 markers as competing methods do, GARD delivers results with 90 % prediction. This can be compared to the gold standard, animal tests on mice, that provide only 72 % prediction. SenzaGen’s test also has the ability to measure potency (strength) of a substance and can thus determine the degree to which a substance is an allergen. Existing, competing tests do not achieve sufficient specificity / sensitivity and lack the ability to measure the potency of a substance.



Ian Kimber joins the board of SenzaGen AB

SenzaGen AB, a diagnostic spin-off company from Lund University, is proud to announce that Prof. Ian Kimber will join the Board of Directors.

SenzaGen AB is a diagnostic company working to replace animal testing for sensitization predictions.

Prof Ian Kimber is currently Professor of Toxicology and Associate Dean for Business Development in the Faculty of Life Sciences at the University of Manchester. He has broad research interests at the interface between toxicology and immunology, with a particular focus on allergy and inflammation.  Professor Kimber holds, and has held, a variety of positions on national and international expert and scientific advisory committees. Currently these include the following:  Member UK Medicines and Healthcare products Regulatory Agency (MHRA) Committee for Safety of Devices, Programme Advisor Food Standards Agency Food Allergy and Intolerance Research Programme, and member MRC Translational Research Group. Professor Kimber was previously President of the British Toxicology Society (BTS) (2012-2014), and Chairman of the Board of the UK National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs) (2008-2013).  

“I am delighted to be joining the Board of SenzaGen at such an exciting period of growth. Since our initial studies that led to the development and ultimate validation of the Local Lymph Node Assay I have maintained a keen interest in chemical allergy and in the development of novel approaches for improved hazard identification and risk assessment. In this respect harnessing fully the opportunities afforded by developments in the biomedical sciences is critical, and the work of SenzaGen in exploiting a genomic approach to pathway analysis and risk assessment is at the cutting edge of new developments”, says Prof Kimber.

Ian Kimber has published over 600 research papers, review articles and book chapters, and serves currently on the editorial boards of toxicology, immunology, dermatology and pathology journals.

“We are very happy to have Prof Kimber on board in SenzaGen”, since he is one of the forerunners in sensitization testing, says Prof Carl Borrebaeck, “his knowledge and interest in the field of allergy and replacement of animal testing with new technologies is very important to us. We value his participation in developing the next generation tests.”

Systematic evaluation of non-animal test methods for skin sensitisation safety assessment.

Toxicology in Vitro   Volume 29, Issue 1, February 2015, Pages 259–270

Kerstin Reisinger, Sebastian Hoffmann, Nathalie Alépée, Takao Ashikaga, Joao Barroso, Cliff Elcombe, Nicola Gellatly, Valentina Galbiati, Susan Gibbs, Hervé Groux, Jalila Hibatallah, Donald Keller, Petra Kern, Martina Klaric, Susanne Kolle, Jochen Kuehnl, Nathalie Lambrechts, Malin Lindstedt, Marion Millet, Silvia Martinozzi-Teissier, Andreas Natsch, Dirk Petersohn, Ian Pike, Hitoshi Sakaguchi, Andreas Schepky, Magalie Tailhardat, Marie Templier, Erwin van Vliet, Gavin Maxwell



The need for non-animal data to assess skin sensitisation properties of substances, especially cosmetics ingredients, has spawned the development of many in vitro methods. As it is widely believed that no single method can provide a solution, the Cosmetics Europe Skin Tolerance Task Force has defined a three-phase framework for the development of a non-animal testing strategy for skin sensitisation potency prediction. The results of the first phase – systematic evaluation of 16 test methods – are presented here. This evaluation involved generation of data on a common set of ten substances in all methods and systematic collation of information including the level of standardisation, existing test data, potential for throughput, transferability and accessibility in cooperation with the test method developers. A workshop was held with the test method developers to review the outcome of this evaluation and to discuss the results. The evaluation informed the prioritisation of test methods for the next phase of the non-animal testing strategy development framework. Ultimately, the testing strategy – combined with bioavailability and skin metabolism data and exposure consideration – is envisaged to allow establishment of a data integration approach for skin sensitisation safety assessment of cosmetic ingredients.


Skin sensitisationTesting strategySafety assessmentNon-animal test methodsAdverse Outcome Pathways

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