The availability of well-documented high-quality clinical pancreas specimens in tissue depositories such as the nPOD allows multiplatform comparative investigations of the biochemical correlates of T1D and T2D disease progression. Mass spectrometry imaging (MSI) is an emerging technique that allows us to determine, in an untargeted manner, the proteins and peptides and monitor their localization within the pancreas. However, its application to specimens preserved using standard formaldehyde-induced crosslinking is limited to molecules that do not have primary amines, and thus excludes endogenous peptides and proteins. Here we are creating a MSI-compatible chemical decrosslinking to enable us to use the tissue banked materials. We found that hydroxylamine hydrochloride in acidic tris(hydroxymethyl)aminomethane reverses the crosslinking between model peptides in vitro. Among amino acid moieties affected by formaldehyde-fixation, arginine, lysine and cysteine demonstrate best performance leading to the restoration of peptide detection in the treated samples. This treatment is compatible with formaldehyde-fixed pancreas and successfully enables the detection of glucagon, insulin and pancreatic polypeptides localized in the α-, β- and γ-cells. Our approach opens important opportunities for multimodal investigation of large sets of well preserved and documented clinical specimens.
D. Lee: None. S. Rubakhin: None. J.V. Sweedler: None.
American Diabetes Association/Pathway to Stop Diabetes (1-18-VSN-19 to J.V.S.)