Quantification of Abscisic Acid in Wheat Leaf Tissue by Direct Enzyme Immunoassay

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Abstract
The plant growth hormone abscisic acid (ABA) modulates plant responses and adaptations to environmental stress. The objective of this study was to develop a procedure which would facilitate broad‐scale screening of wheat (Triticum aestivum L. em. Thell.) germplasm in which ABA content or rate of accumulation in leaf tissue is the selection criterion. The effects of extraction time, C18 reverse‐phase chromatography, solvent volatilization, and dilution of sample extract on quantification of ABA by direct enzyme immunoassay (EIA) were investigated. Results showed that complete volatilization of extraction solvent did not affect ABA determination. Extraction of ABA from wheat leaves, using 800 mL L−1 methanol (MeOH), was complete after 24 h. Interference to EIA due to MeOH was eliminated when MeOH was diluted to ≤80 mL L−1. No apparent inhibitors to EIA were detected by a parallelism test of dilution curves or by partial purification of leaf extract using C18 reverse‐phase chromatography. Simplified and effective procedures are proposed which facilitate the preparation of a large number of leaf samples at minimal expense and labor without sacrificing accuracy of ABA estimation.

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