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On-tissue identification of insulin: in situ reduction coupled with mass spectrometry imaging
Shamina M Green-Mitchell1, Lisa H Cazares, O John Semmes
1Department of Internal Medicine, Eastern Virginia Medical School, Norfolk, VA, USA.
Purpose:
The aim of this study was to use on-tissue reduction followed by MALDI-MS imaging (MSI) to identify an m/z 5812.85 peak, which is over-expressed in healthy human pancreatic tissue compared with type one Diabetes (T1D) tissue.
Experimental Design:
A major constraint of MALDI-MSI is identification of compounds with m/z ≥4000. On-tissue reduction using tris (2-carboxyethyl) phosphine (TCEP) breaks the inter-domain disulphide bonds generating low-molecular-weight peptides amenable to direct MS/MS analysis. Pancreatic tissues from healthy (n=4) and diabetic subjects (n=4) were profiled by MALDI-MSI with/without reduction.
Results:
On-tissue reduction resulted in the loss of the over-expressed 5812.85 m/z peak and the simultaneous appearance of a 3430.664 m/z peak in healthy tissues. The latter peak presumably derived from the 5812.85 m/z peak was identified as the insulin B chain by MS/MS. MALDI-MSI images show that both the 5812.85 insulin peak before reduction and the 3430.664 peak after reduction co-localized with the healthy pancreatic islets.
Conclusion And Clinical Relevance:
On-tissue reduction followed by MALDI-MSI resulted in the identification of insulin and localization of pancreatic islets of langerhans. The approach will be useful in the future identification of novel therapeutic molecular targets to β-cells lost during type one diabetes.
