Development of antibodies against secoisolanciresinol - Application to the immunolocalization of lignans in Linum usitatissimum seeds

Lignans are widely distributed plant metabolites associated with a large range of biological activities In order to gain insight into their biosynthesis and their spatio-temporal accumulation an immunological probe was developed Secondary metabolites generally have too small molecular weight to be antigenic and have to be associated with a carrier protein Secoisolariciresinol was chosen as the hapten and was linked to bovine serum albumin via a spacer arm the p-aminohippuric acid The artificial antigen was injected to New Zealand rabbits The successful production of polyclonal antibodies against secolsolariciresinol was assessed with indirect enzyme immunosorbent assay (ELISA) by comparison with pre-immune serum and by competitive assays using dilutions of secoisolariciresinol standards The antibodies had an IC50 value of 94 mu g/ml and showed moderate cross-reactivities with structurally related compounds They were thus used to immunolocalize lignans in flaxseed (Linum usitatissimum) one of the richest sources of lignans The immunohistochemical labeling allowed us to localize for the first time lignans in planta They are mainly localized in the secondary wall of the sclerite cells of the outer integument of the seed A very light labeling is also observed in cytoplasmic inclusions of the endosperm The results were correlated with HPLC analytical results which enabled to evaluate the relative lignan quantities in flaxseed about 90% of the metabolites are localized in the outer integument (C) 2010 Elsevier Ltd All rights reserved