Immunoaffinity co-purification of cytokinins and analysis by high-performance liquid chromatography with ultraviolet-spectrum detection

Planta
Björn NicanderElisabeth Tillberg

Abstract

A rapid methodology for the simultaneous analysis of a large number of cytokinins is presented. The cross-reactivity of a mixture of polyclonal antibodies against zeatin riboside and isopentenyladenosine was exploited in a protocol that can be used for immunoaffinity purification of 23 additional cytokinins. Ligands include the cytokinin bases zeatin, dihydrozeatin, isopentenyladenine, benzyl-adenine and kinetin, and their corresponding nucleoside, nucleoside-5'-monophosphate, and 9-glucoside derivatives, as well as cis-zeatin, cis-zeatin riboside, the 2-methylthiol derivatives of isopentenyladenosine and zeatin riboside, and benzyl-adenine-3-glucoside. Mixtures of cytokinins could be retained with high recoveries of all the components. Immunoaffinity purification of extracts of Arabidopsis thaliana (L.) Heynh. and Solarium tuberosum L. gave fractions clean enough, as verified by gas chromatographymass spectrometry (GC-MS), to allow analysis of endogenous cytokinins using a single high-performance liquid chromatography (HPLC) step with on-line UV-spectrum detection. The detection limit was 4-6 pmol. The procedure described forms a routine assaying technique that is faster and simpler, yet yields better qualitative and quantitat...Continue Reading

Citations

Jun 17, 1998·The Plant Journal : for Cell and Molecular Biology·Hitoshi SakakibaraT Sugiyama
Nov 12, 2013·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Bao-Dong CaiYu-Qi Feng
Mar 29, 2014·Planta·Danuse TarkowskaM Strnad
Apr 25, 2001·Physiologia Plantarum·Huaibi ZhangPaula Elizabeth Jameson
Oct 27, 2006·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Veronika HradeckáM Strnad
Sep 30, 2005·Journal of Chromatography. a·Eva HauserováM Strnad
Jan 17, 2004·The Journal of Biological Chemistry·Hiroyuki KasaharaHitoshi Sakakibara

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