Validation of antibody-based tools for galanin research

•Most commercial galanin and galanin receptor antibodies are not specific.•Four antibodies against the human galanin system are validated.•The specificity of some commercially available galanin ELISA kits is questionable.•A protocol for development of a specific human galanin ELISA is established.•Galanin can be measured directly in plasma using the in-house ELISA. Antibodies are an integral biomedical tool, not only for research but also as therapeutic agents. However, progress can only be made with sensitive and specific antibodies. The regulatory (neuro)peptide galanin and its three endogenous receptors (GAL1-3-R) are widely distributed in the central and peripheral nervous systems, and in peripheral non-neuronal tissues. The galanin system has multiple biological functions, including feeding behavior, pain processing, nerve regeneration and inflammation, to name only a few. Galanin could serve as biomarker in these processes, and therefore its receptors are potential drug targets for various diseases. For that reason, it is of paramount interest to precisely measure galanin peptide levels in tissues and to determine the cellular and subcellular localization of galanin receptors. A plethora of antibodies and antibody-based tools, including radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) kits, are commercially available to detect galanin and its receptors. However, many of them lack rigorous validation which casts doubt on their specificity. A goal of the present study was to raise awareness of the importance of validation of antibodies and antibody-based tools, with a specific focus on the galanin system. To that end, we tested and report here about commercially available antibodies against galanin and galanin receptors that appear specific to us. Furthermore, we investigated the validity of commercially available galanin ELISA kits. As the tested ELISAs failed to meet the validation requirements, we developed and validated a specific sandwich ELISA which can be used to detect full-length galanin in human plasma.