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Measurement of human latent transforming growth factor beta 1 using a latency associated protein reactive elisa
Stockholms universitet, Naturvetenskapliga fakulteten, Wenner-Grens institut, Avdelningen för immunologi.
2012 (Engelska)Ingår i: JIM - Journal of Immunological Methods, ISSN 0022-1759, E-ISSN 1872-7905, Vol. 379, nr 1-2, s. 23-29Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Human Transforming Growth Factor (TGF)-beta 1, one of three TGF-beta isoforms, is a pleotropic cytokine critical for many physiological and immunological processes. TGF-beta 1 is secreted in a latent form, linked to Latency Associated Protein (LAP). Analysis of Latent TGF-beta 1 by TGF-ELISA requires dissociation of TGF-beta 1 from LAP, e.g. by acidification of samples. The ELISA then measures total TGF-beta 1, equivalent to dissociated Latent TGF-beta 1 plus any free TGF-beta 1 present prior to acidification. Evolutionary conservation of TGF-beta 1 across mammals also renders TGF-beta 1 ELISAs reactive with TGF-beta 1 in bovine serum often used in human cell cultures. To enable a direct analysis of Latent TGF-beta 1, monoclonal antibodies were made against LAP from human latent TGF-beta 1 and used to develop a LAP ELISA detecting Latent TGF-beta 1. The ELISA did not react with LAP from human Latent TGF-beta 2 or 3, respectively, nor with Latent TGF-beta in bovine serum. EDTA-containing plasma from healthy subjects (n = 20) was analyzed by conventional TGF-beta 1 ELISA and LAP ELISA. By TGF-beta 1 ELISA, total TGF-beta 1 were detected in all samples (median 133 pM, range 34-348 pM); low levels of free TGF-beta 1 found in 8/20 non-addified samples showed that >98.5% of the total TGF-beta 1 derived from Latent TGF-beta 1. Latent TGF-beta 1 found in non-acidified samples by LAP ELISA (median 154 pM, range 48-403 pM) was comparable in molar levels to, and correlated with, total TGF-beta 1 (r(s) 0.96, p<0.0001). A similar agreement between the total TGF-beta 1 and the LAP ELISA was found with citrate- and heparin-containing plasma. The LAP ELISA facilitates analysis of Latent TGF-beta 1 without sample acidification and is not compromised by the presence of bovine serum in human cell supernatants.

Ort, förlag, år, upplaga, sidor
2012. Vol. 379, nr 1-2, s. 23-29
Nyckelord [en]
ELISA, Cytokine, TGF-beta 1, Latent TGF-beta 1, Total TGF-beta 1
Nationell ämneskategori
Immunologi Biokemi och molekylärbiologi
Identifikatorer
URN: urn:nbn:se:su:diva-79768DOI: 10.1016/j.jim.2012.02.016ISI: 000304019600003OAI: oai:DiVA.org:su-79768DiVA, id: diva2:551886
Anmärkning

AuthorCount:4;

Tillgänglig från: 2012-09-12 Skapad: 2012-09-11 Senast uppdaterad: 2017-12-07Bibliografiskt granskad

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JIM - Journal of Immunological Methods
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