title: Towards a human proteome atlas: high-throughput generation of mono-specific antibodies for tissue profiling.
abstract: A great need exists for the systematic generation of specific antibodies to explore the human proteome. Here, we show that antibodies specific to human proteins can be generated in a high-throughput manner involving stringent affinity purification using recombinant protein epitope signature tags (PrESTs) as immunogens and affinity-ligands. The specificity of the generated affinity reagents, here called mono-specific antibodies (msAb), were validated with a novel protein microarray assay. The success rate for 464 antibodies generated towards human proteins was more than 90% as judged by the protein array assay. The antibodies were used for parallel profiling of patient biopsies using tissue microarrays generated from 48 human tissues. Comparative analysis with well-characterized monoclonal antibodies showed identical or similar specificity and expression patterns. The results suggest that a comprehensive atlas containing extensive protein expression and subcellular localization data of the human proteome can be generated in an efficient manner with mono-specific antibodies.
authors.affiliation: Department of Biotechnology, Royal Institute of Technology (KTH), Stockholm, Sweden.
authors.forename: Peter
authors.initials: P
authors.lastname: Nilsson
authors.forename: Linda
authors.initials: L
authors.lastname: Paavilainen
authors.forename: Karin
authors.initials: K
authors.lastname: Larsson
authors.forename: Jenny
authors.initials: J
authors.lastname: Odling
authors.forename: Mårten
authors.initials: M
authors.lastname: Sundberg
authors.forename: Ann-Catrin
authors.initials: AC
authors.lastname: Andersson
authors.forename: Caroline
authors.initials: C
authors.lastname: Kampf
authors.forename: Anja
authors.initials: A
authors.lastname: Persson
authors.forename: Cristina
authors.initials: C
authors.lastname: Al-Khalili Szigyarto
authors.forename: Jenny
authors.initials: J
authors.lastname: Ottosson
authors.forename: Erik
authors.initials: E
authors.lastname: Björling
authors.forename: Sophia
authors.initials: S
authors.lastname: Hober
authors.forename: Henrik
authors.initials: H
authors.lastname: Wernérus
authors.forename: Kenneth
authors.initials: K
authors.lastname: Wester
authors.forename: Fredrik
authors.initials: F
authors.lastname: Pontén
authors.forename: Mathias
authors.initials: M
authors.lastname: Uhlen
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry* / Q000302:isolation & purification; D000918:Antibody Specificity; D002846:Chromatography, Affinity; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D040081:Protein Array Analysis; D020543:Proteome / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology
chemical_list: D000906:Antibodies; D020543:Proteome; D011994:Recombinant Proteins