A Genecentric Human Protein Atlas for Expression Profiles Based on Antibodies

Abstract

An attractive path forward in proteomics is to experimentally annotate the human protein complement of the genome in a genecentric manner. Using antibodies, it might be possible to design protein-specific probes for a representative protein from every protein-coding gene and to subsequently use the antibodies for systematical analysis of cellular distribution and subcellular localization of proteins in normal and disease tissues. A new version (4.0) of the Human Protein Atlas has been developed in a genecentric manner with the inclusion of all human genes and splice variants predicted from genome efforts together with a visualization of each protein with characteristics such as predicted membrane regions, signal peptide, and protein domains and new plots showing the uniqueness (sequence similarity) of every fraction of each protein toward all other human proteins. The new version is based on tissue profiles generated from 6120 antibodies with more than five million immunohistochemistry-based images covering 5067 human genes, corresponding to ϳ25% of the human genome. Version 4.0 includes a putative list of members in various protein classes, both functional classes, such as kinases, transcription factors, G-protein-coupled receptors, etc., and project-related classes, such as candidate genes for cancer or cardiovascular diseases. The exact antigen sequence for the internally generated antibodies has also been released together with a visualization of the application-specific validation performed for each antibody, including a protein array assay, Western blot analysis, immunohistochemistry, and, for a large fraction, immunofluorescencebased confocal microscopy. New search functionalities have been added to allow complex queries regarding protein expression profiles, protein classes, and chromosome location. The new version of the protein atlas thus is a resource for many areas of biomedical research, including protein science and biomarker discovery. Molecular

Reproducibility crisis: Blame it on the antibodies

“…Perhaps the largest assessment comes from work published by the Human Protein Atlas, a Swedish consortium that aims to generate antibodies for every protein in the human genome. It has looked at some 20,000 commercial antibodies so far and found that less than 50% can be used effectively to look at protein distribution in preserved slices of tissue 5 . This has led some scientists to claim that up to half of all commercially available antibodies are unreliable.…”

Reproducibility: Standardize antibodies used in research

“…Alarmingly, there are serious flaws in the reliability of antibodies, the most widely used class of protein-binding reagent 2,3 .…”

“…In the lab, biologists have long used them to track proteins of interest because they bind to specific targets. But in a 2008 study 3 , fewer than half of around 6,000 routinely used commercial antibodies recognized only their specified targets, with some manufacturers producing consistently good antibodies, and others consistently poor ones.…”

Novel imaging tools for investigating the role of immune signalling in the brain

“…Current estimates suggest approximately 50 per cent of antibodies are either unreactive towards the purported epitope or cross-react with others (Berglund et al 2008). This problem is amplified by common chemical treatments on the collected tissues that are necessary for many immunohistochemical techniques.…”