Avtor/Urednik		Wozniak, Gordana; Obermayr, Eva; Jeras, Matjaž; Knežević, Mio; Rueker, Florian
Naslov		An ELISA for the detection of TIMP-1 based on recombinant single chain Fv fusion proteins
Tip		članek
Vir		Clin Chim Acta
Vol. in št.		Letnik 335, št. 1-2
Leto izdaje		2003
Obseg		str. 49-57
Jezik		eng
Abstrakt		Background: Altered serum levels of TIMP-1 are an indicator of various pathological states. To quantitate TIMP-1 in biological samples, we have previously isolated TIMP-1 specific single-chain Fv fragments (scFvs) using phage-display. In the work presented here, we have used these scFvs to establish a TIMP-1 ELISA based exclusively on recombinant scFv fusion proteins. Methods: Two distinct TIMP-1 specific scFvs were used as the antigen binding components after being genetically fused to the N-termini of two different fusion protein constructs. One fusion protein, comprising a C(L) domain, serves as a coating reagent, while the second fusion protein with a modified form of bacterial alkaline phosphatase is used as a detection reagent. A double antibody sandwich-ELISA was then established and optimized. Results: An ELISA for the quantitation of tissue inhibitor of metalloproteinase (TIMP)-1, based entirely on recombinant antibody fragments, was developed as an alternative to assays using polyclonal antisera or monoclonal antibodies. Its performance was shown to compare well with a conventional ELISA. Finally, TIMP-1 concentrations in the sera of sixty healthy individuals were determined. Conclusion: The assay described here provides a standardized, reliable and readily available means of quantitation of TIMP-1 in a large number of blood samples.
Deskriptorji		ENZYME-LINKED IMMUNOSORBENT ASSAY METALLOPROTEINASES ALKALINE PHOSPHATASE IMMUNOGLOBULIN FRAGMENTS RECOMBINANT FUSION PROTEINS ESCHERICHIA COLI BLOTTING, WESTERN