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All assays through which the antibody has been validated. Assays&annotation provide a detailed description of the different assays. The pie-charts indicate degree of validation.
Immunohistochemistry is used for validating antibody reliability by assessing staining pattern in 44 normal tissues. Validation scores include Enhanced, Supported, Approved and Uncertain.
Results of validation by standard or enhanced validation based on assessment of antibody performance in 44 normal tissues.
Standard validation results in scores Supported, Approved or Uncertain. An image representative of the antibody staining pattern is shown.
Enhanced validation results in the score Enhanced and includes two methods: Orthogonal validation and Independent antibody validation. For orthogonal validation, representative images of high and low expression are shown. For independent antibody validation, four images of each independent antibody are displayed.
Immunohistochemistry is used for validating antibody reliability by assessing staining pattern in 44 normal tissues. Validation scores include Enhanced, Supported, Approved and Uncertain.
Immunohistochemical staining of human gallbladder shows strong cytoplasmic positivity in glandular cells.
Retrievali
Antigen retrieval is a method used to restore/retrieve the epitope (antibody bidning region) of the target protein, cross-linked, and thus masked, during tissue preserving fixative treatment of the tissues.
Conformance of the expression pattern with available gene/protein characterization data in scientific literature and data from bioinformatic predictions.
UniProt is used as the main source of gene/protein characterization data and when relevant, available publications and other sources of information are researched in depth. Extensive or sufficient gene/protein data requires that there is evidence of existence on a protein level and that a substantial quantity of published experimental data is available from literature and public databases. Limited protein/gene characterization data does not require evidence of existence on a protein level and refers to genes for which only bioinformatic predictions and scarce published experimental data is available.
Partly consistent with extensive gene/protein characterization data.
RNA consistencyi
Consistency between immunohistochemistry data and internally generated RNA-seq data is divided into five different categoreies: i) Consistent with RNA expression data, ii) Mainly consistent with RNA expression data, iii) Mainly not consistent with RNA expression data, iv) Not consistent with RNA expression data, and v) No internal RNA expression data available for correlation.
No internal RNA expression data available for correlation.
WESTERN BLOTi
A Western blot analysis is performed on a panel of human tissues and cell lines to evaluate antibody specificity. For antibodies with unreliable result a revalidation using an over-expression lysate is performed.
Western Blot is used for quality control of the polyclonal antibodies generated in the project. After purification, the antibodies are used to detect bands in a setup of lysate and different tissues. The result is then scored Enhanced, Supported, Approved, or Uncertain.
Enhanced validation includes five different methods: Genetic validation, Recombinant expression validation, Independent antibody validation, Orthogonal validation and Capture MS validation.
The staining of an antibody is evaluated by Western Blot through analysis of samples from different cell lysates. A supportive score is given if band(s) of predicted size in kDa (+/-20%) is detected.
Only bands not corresponding to the predicted size. Analysis performed using a standard panel of samples.
The Human Protein Atlas project is funded
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