Immunoaffinity chromatography of abscisic acid combined with electrospray liquid chromatography-mass spectrometry.
Polyclonal antibodies with high specificity for C1-immobilised (+)-cis,trans-abscisic acid (ABA) were raised, characterised by enzyme-linked immunosorbent assay (ELISA) and used for preparation of an immunoaffinity chromatography (IAC) gel. The detection limit of the ELISA was approximately 4.6x10(-10)mol/L. Sensitive electrospray liquid chromatography-mass spectrometry (LC-ESI-MS) methods were also developed with detection limits below 0.1x10(-12)mol. The IAC allowed quick, single-step processing of samples prior to the analyses. The LC-ESI-MS and LC-ELISA techniques were used for comparative estimation of endogenous ABA levels in immunoaffinity purified extracts of normal and water-stressed Nicotiana tabacum L. leaves. The analytical approaches were validated using deuterium- and tritium-labelled internal standards, respectively. The IAC method was found to be highly effective, sensitive and convenient for isolating the target analyte from plant material.
Hradecká
Veronika
V
Laboratory of Growth Regulators, Palacký University & Institute of Experimental Botany ASCR, Slechtitelů 11, CZ-783 71 Olomouc, Czech Republic. hradeckav@post.cz
Novák
Ondrej
O
Havlícek
Libor
L
Strnad
Miroslav
M
D000040:Abscisic Acid / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D002846:Chromatography, Affinity / Q000379:methods*; D002853:Chromatography, Liquid / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D018515:Plant Leaves / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014026:Nicotiana / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D014867:Water / Q000378:metabolism / Q000494:pharmacology
D014867:Water; D000040:Abscisic Acid