Monoclonal Antibody-Based Enzyme Linked Immunosorbent Assay for the Analysis of Jasmonates in Plants

摘要

Methyl jasmonate (MeJA) and its free-acid form,jasmonic acid (JA) are naturally occurring plant growth regulators widely distributed in higher plants.In order to improve the sensitivity for the analysis of MeJA at low levels in small amounts of plant samples,a monoclonal antibody (MAb) (designated as MAb 3E5D7C4B6) against MeJA was derived from a JAbovine serum albumin (BSA) conjugate as an immunogen.The antibody belongs to the IgG1 subclass with a κ type light chain and has a dissociation constant of approximately 6.07 x 10-9 M.MAb3E5D7C4B6 is very specific to MeJA.It was used to develop a direct competitive enzyme-linked immunosorbent assay (dcELISA),conventional and simplified indirect competitive ELISAs (icELISA).JA was derivatized into MeJA for the ELISA analysis.The IC50 value and detection range for MeJA were,respectively,34 and 4-257 ng/mL by the conventional icELISA,21 and 3-226 ng/mL by the simplified icELISA and 5.0 and 0.7-97.0 ng/mL by the dcELISA.The dcELISA was more sensitive than either the conventional or simplified icELISA.The assays were used to measure the content of jasmonates as MeJA in tobacco leaves under drought stress or inoculated with tobacco mosaic virus and tomato leaves inoculated with tomato mosaic virus or Lirioinyza sativae Blanchard as compared with the corresponding healthy leaves.The increased jasmonates content indicated its role in response to the drought stress and pathogens.