Development of highly fluorescent detection reagents for the construction of ultrasensitive immunoassays

Analytical Chemistry
Q P QinK Pettersson

Abstract

We developed two kinds of highly fluorescent streptavidin-based conjugates for use as universal detection reagents in ultrasensitive immunoassays. The direct conjugate was produced by covalently linking streptavidin to poly(Glu: Lys) which was labeled heavily with Eu chelates; the indirect conjugate was made by first conjugating bovine serum albumin (BSA) to poly(Glu:Lys) labeled heavily with Eu chelates and then further linking streptavidin to the conjugate of BSA-poly(Glu:Lys)-Eu chelate. Both direct and indirect conjugates were used to construct a highly sensitive time-resolved fluorometric assay for prostate-specific antigen (PSA). Of two monoclonal antibodies used in the assay, one was coated on the well surface of the microtitration strips, and the other was biotinylated. When 10 microL of sample volume was used, we found that the assay using the indirect conjugate had a detection limit of 0.006 microg/L, which was approximately 5.6-fold more sensitive than the one using Eu chelate directly labeled detection antibody and 6.8-fold more sensitive than the one using Eu chelate-labeled streptavidin. However, the assay that used the direct conjugate was 1.5-fold more sensitive than the one that utilized the indirect conjugate....Continue Reading

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Citations

Jan 6, 2006·Analytical and Bioanalytical Chemistry·King-Keung SinReinhard Renneberg
Sep 17, 2005·Journal of Fluorescence·I Hemmilä, V Laitala
Sep 17, 2005·Journal of Fluorescence·Jingli Yuan, Guilan Wang
Feb 1, 2003·Clinica Chimica Acta; International Journal of Clinical Chemistry·Tero SoukkaTimo Lövgren
Nov 1, 2006·Journal of Nanobiotechnology·Chitta Ranjan PatraDebabrata Mukhopadhyay
Jun 30, 2005·Analytical Sciences : the International Journal of the Japan Society for Analytical Chemistry·Lihua PanMin Lin
Jul 22, 2004·Analytical Biochemistry·Lasse VälimaaTimo Lövgren
Jun 30, 2006·Clinical Biochemistry·Johanna YlikotilaKim Pettersson
Jan 12, 2007·Molecular BioSystems·Uwe R Müller
Sep 21, 2011·Dalton Transactions : an International Journal of Inorganic Chemistry·Weili JiangChunhui Huang
Aug 1, 2013·ChemPlusChem·Gang YuChunhui Huang
Apr 15, 2004·Journal of the American Chemical Society·Nicolas WeibelRaymond Ziessel
Oct 15, 2011·The Journal of Physical Chemistry. B·Qiang Li, Stefan Seeger

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