Antibody Validation

At Atlas Antibodies, we work extensively with antibody validation to produce specific, selective, and reproducible antibodies you can trust. 

All our antibodies are rigorously evaluated for specificity and performance and are characterized in several applications. In addition, an extra layer of validation, Enhanced Validation, is performed following the recommendations of the International Working Group for Antibody Validation (IWGAV) published in Nature Methods.

Continue reading to learn about Enhanced Validation and how we employed it to ensure the specificity of our antibodies. 

Characterization and Validation of Antibodies

Antigen Design

For our antibodies, specificity starts with the design. The antibodies are developed against recombinant human Protein Epitope Signature Tags (PrESTs) of approximately 50 to 150 amino acids. The PrEST sequences are designed to have the lowest possible identity to other human proteins. 

Protein Array

The specificity and purity are validated by protein array against 364 human recombinant protein fragments.

Functional validation and characterization

The Triple A Polyclonals and PrecisA Monoclonals manufactured by Atlas Antibodies are designed, characterized, validated, and approved in IHC, ICC-IF, and WB. For IHC, each antibody is evaluated in 44 normal tissues and the 20 most common cancers. 

Each antibody is accompanied by 500 staining images, manually annotated by researchers and pathologists, and available on the Human Protein Atlas website. 

Antibody approval

Antibody specificity is further evaluated considering the literature, bioinformatics, and RNA sequencing data.

Transparency

We believe that transparency and open access are crucial for research. Therefore, all characterization data, including images, are freely available on the Atlas Antibodies and the Human Protein Atlas websites. In addition, we provide you with the exact immunogen sequence we used to design our antibodies. 

Enhanced Validation

On top of the extensive validation and characterization always performed for our antibodies, we perform application-specific Enhanced validation. The enhanced validation follows the guidelines proposed by the International Working Group for Antibody Validation (IWGAV) in Nature Methods.

The working group was formed by distinguished researchers worldwide to "formulate the best approaches for validating antibodies used in common research applications and to provide guidelines that ensure antibody reproducibility." 1 IWGAV proposed five conceptual pillars for antibody validation to be used in an application-specific manner. At least one of the five pillars must be used to validate an antibody in a specific application. At Atlas Antibodies, we base the Enhanced Validation on Human Protein Atlas' interpretation of these pillars, as explained here.

Learn about Enhanced Validation

Learn about Enhanced Validation

Enhanced validation offers increased security of antibody specificity in a defined context. This is ensured by using the most relevant validation method for each combination of protein, sample, and application. 

Learn more

Reproducibility

In addition to performing a thorough validation, it is vital to ensure antibody reproducibility. This means that a new antibody lot must perform equally compared to the previous lot so that every experiment can be repeated with the same results. Therefore, at Atlas Antibodies, we test all new antibody lots in parallel to existing lots in a large set of samples.

 

1 The group of distinguished researchers from Stanford University, Yale University, MIT, UCSD, and NIH in the USA, University of Toronto in Canada, the EMBL in Germany, Niigata University in Japan and the Science for Life Laboratory in Sweden, was formed to develop formal guidelines for antibody validation and reproducibility to be used by antibody manufacturers to validate antibodies in a standardized manner (Nature Methods, September 2016).

 

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