Abstract
A new monoclonal antibody (mAb) was generated against abscisic acid (ABA), and an indirect enzyme-linked immunosorbent assay (ELISA) using this mAb was developed for convenient quantitative analysis of ABA levels in rice leaf extracts. The mAb, raised against (+-)-ABA conjugated to bovine serum albumin (BSA) through its carboxyl group (C1), reacted preferentially with the (+)-ABA enantiomer, and equally well with both free and methyl-ester (+-)-ABA. Cross-reactivity with several ABA-related compounds was negligible. Linearity was obtained between 3 and 1000 pmo1 of (+)-ABA. The ABA-mAb was further used to quantitate pmol quantities of (+)-ABA in attached and detached rice leaves. Results obtained with such ELISA quantitation showed an increase in the free ABA content of detached rice leaves at progressive stages of senescence, which was regarded as a senescence-related response. This quantitation compared favorably with other presently used techniques for ABA determination, with regard to their detection limits, cost and assay time. The results suggest that the combination of a specific mAb with a sensitive ELISA technique is quite promising for quantitation of ABA.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Aharoni N and Richmond AE (1978) Endogenous gibberellin and abscisic acid content as related to senescence of detached lettuce leaves. Plant Physiol 62: 224–228
Belefant H and Fong F (1989) Abscistic acid ELISA: organic acid interference. Plant Physiol 91: 1467–1470
Biswas AK and Choudhuri MA (1980) Mechanism of monocarpic senescence in rice. Plant Physiol. 65:340–345
Daie J and Wyse R (1982) Adaptation of the enzyme-linked immunosorbent assay (ELISA) to the quantitative analysis of abscisic acid. Anal Biochem 119: 365–371
Galfre G and Milstein C (1981) Preparation of monoclonal antibodies: strategies and procedures. Methods Enzymol 73: 3–46
Gepstein S and Thimann KV (1980) Changes in the abscisic acid content of oat leaves during senescence. Proc Natl Acad Sci USA 77: 2050–2053
Hadas E and Theilen G (1987) Production of monoclonal antibodies: The effect of hybridoma concentration on the yield of antibody-producing clones. J Immunol Methods 96: 3–6
Harris MJ and Dugger WM (1986) The occurrence of abscisic acid and abscisyl-β-D-glucopyranoside in developing and mature citrus fruit as determined by enzyme immunoassay. Plant Physiol. 82: 339–345
Harris MJ, Outlaw WH Jr, Mertens Rand Weiler EM (1988) Water-stress-induced changes in the abscisic acid content or guard cells and other cells of Vicia faba L. leaves as determined by enzyme-amplified immunoassay. Proc Natl Acad Sci USA 85: 2584–2588
Henson IE (1985) Modification of leaf size in rice (Oryza sativa L.) and its effects on water stress-induced abscisic acid accumulation. Annals Bot 56: 481–487
Hubick KT and Reid DM (1980) A rapid method of extraction and analysis of abscisic acid from plant tissue. Plant Physiol 65: 523–525
Kao CH (1977) Senescence of rice leaves I. Comparison between detached and intact leaves. Memoirs Coll Agric Natl Taiwan Univ 17: 98–102
Kao CH and Yang SF (1982) Light inhibition of the conversion of 1-aminocyclopropane-1-caboxylic acid to ethylene in leaves is mediated through carbon dioxide. Planta 155: 261–266
Leroux B, Maldiney R, Miginiac E, Sossountzov L and Sotta B (1985) Comparative quantitation of abscisic acid in plant extracts by gas-liquid chromatography and an enzyme-linked immunosorbent assay using avidin-biotin system. Planta 166: 524–529
Mertens, R, Deus-Neumann B and Weiler EW (1983) Monoclonal antibodies for the detection and quantitation of the endogenous plant growth regulator, abscisic acid. FEBS Letters 160: 269–272
Norman SM, Poling SM and Maier VP (1988) An indirect enzyme-linked immunosorbent assay for (+)-abscisic acid in Citrus, Ricinus and Xanthium leaves. J Agric Food Chem 36: 225–231
Pekic S and Quarrie SA (1988) Abscisic acid in leaves of inbred lines and F1 hybrids of maize growing in the field and its relationship to diurnal and seasonal trends in leaf water potential. Annals Bot 61: 669–678.
Quarrie SA, Whitford PN, Appleford NEJ, Want TL, Cook SK, Henson IE and Loveys BR (1988) A monoclonal antibody to (S)-abscisic acid: its characterization and use in a radioimmunoassay for measuring abscisic acid in crude extracts of cereal and lupin leaves. Planta 173: 330–339
Ross GS, Elder PA, McWha JA, Pearce D and Pharis RP (1987) The development of an indirect enzyme linked immunoassay for abscisic acid. Plant Physiol 85: 46–50
Tijssen P (1986) Quantitative enzyme immunoassay techniques. In: RH Burdon, PH Knippenberg, eds. Practice and Theory of Enzyme Immunoassays, 344–384. The Netherlands: Elsevier
Vernieri P, Perata P, Armellini D, Bugnoli M, Presentini R, Lorenzi R, Ceccarelli N, Alpi A and Tognoni F (1989) Solid phase radioimmunoassay for the quantitation of abscisic acid in plant crude extracts using a ne7w monoclonal antibody. J Plant Physiol 134: 441–446
Walker-Simmons MK, Reaney MJT, Quarrie SA, Perata P, Vernieri P and Abrams SR (1991) Monoclonal antibody recognition of abscisic acid analogs. Plant Physiol 95: 46–51
Walton DC (1980) Biochemistry and physiology of abscisic acid. Annu Rev Plant Physiol 31: 453–489
Weiler EW (1980) Radioimmunoassays for the differential and direct analysis of free and conjugated abscisic acid in plant extracts. Planta 148: 262–272
Weiler EW (1982) An enzyme-immunoassay for cis-(+)abscisic acid. Physiol Plant 54: 510–514
Weiler EW (1984) Immunoassay of plant growth regulators. Annu Rev Plant Physiol 35: 85–95
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Philosoph-Hadas, S., Hadas, E. & Aharoni, N. Characterization and use in ELISA of a new monoclonal antibody for quantitation of abscisic acid in senescing rice leaves. Plant Growth Regul 12, 71–78 (1993). https://doi.org/10.1007/BF00144585
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00144585