Anti-l-light chain-peptide antibodies are suitable for the immunohistochemical classification of AL amyloid

Abstract

We aimed to test whether antibodies raised against recombinant peptides corresponding to the variable region of immunoglobulin light chains are suitable for the immunohistochemical classification of amyloid. The Entrez database of the National Center for Biotechnology Information (NCBI) was searched for all protein sequence entries which met the search criteria “amyloid” and “lambda light chain”. Sixty-four different l-light chain-derived amyloid protein sequences were retrieved, aligned and categorized into the V region subgroups of l-light chain detailed by the NCBI, i.e. subgroup I (21 protein sequences), II (14), III (6), IV (1), V (1) and VI (21). V region subgroup I was chosen for epitope sequence selection and two rabbits were immunized with the following peptides: NH2- ISCSGSSSNIGSNTV-CONH2 and NH2-QRPSG VPDRFSGSKSGTS-CONH2. Sensitivity and specificity of the IgG-purified antibodies was tested by Western blotting using amyloid A- (AA), ALl- and ALk-amyloid proteins, and by immunohistochemistry on tissue microarrays with 110 different amyloid containing tissue samples obtained at autopsy from 22 patients, and on 27 biopsy specimens from a series of 24 patients. Our peptide antibodies specifically stained AL amyloid l- light chain-origin, in both Western blots and formalinthat peptide-antibodies directed against immunoglobulin- derived l-light chain proteins can be applied for the immunohistochemical classification of amyloid. This offers the opportunity to generate a large set of anti- l-light chain protein-antibodies for the immunohistochemical classification of amyloid independently from native human tissue sources.