title: On-tissue identification of insulin: in situ reduction coupled with mass spectrometry imaging.
abstract: OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Internal Medicine, Eastern Virginia Medical School, Norfolk, VA, USA.
authors.forename: Shamina M
authors.initials: SM
authors.lastname: Green-Mitchell
authors.forename: Lisa H
authors.initials: LH
authors.lastname: Cazares
authors.forename: O John
authors.initials: OJ
authors.lastname: Semmes
authors.forename: Jerry L
authors.initials: JL
authors.lastname: Nadler
authors.forename: Julius O
authors.initials: JO
authors.lastname: Nyalwidhe
mesh_terms: D016022:Case-Control Studies; D003922:Diabetes Mellitus, Type 1 / Q000175:diagnosis* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007328:Insulin / Q000032:analysis / Q000737:chemistry; D007515:Islets of Langerhans / Q000033:anatomy & histology / Q000378:metabolism*; D057054:Molecular Imaging / Q000379:methods*; D010084:Oxidation-Reduction; D010455:Peptides / Q000032:analysis*; D010720:Phosphines / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry
chemical_list: D007328:Insulin; D010455:Peptides; D010720:Phosphines