Evaluation of the SELDI‐TOF MS technique for protein profiling of pancreatic islets exposed to glucose and oleate

Abstract

The aim of the study was to evaluate the SELDI-TOF MS technique for pancreatic islet research. Mouse islets were cultured at low or high glucose levels in the absence or presence of oleate and characterized by measuring insulin secretion and oxygen tension. Subsequently, the islets were protein profiled. Up to 200 different peaks could be detected in a single experiment with the majority of peaks corresponding to proteins with masses below 30 kDa. By combining different protein arrays, the number of detected peaks could be increased further. The optimal binding of islet proteins was achieved using the anionic exchange array and phosphate buffer (pH 6) when the binding of insulin was low, which allowed other less abundant proteins to be captured. When islets from different culture conditions were profiled and analyzed, in total 25 proteins were found to be oleate/glucose-regulated. An oleate-regulated protein was chosen for identification work, which was conducted by passive elution from SDS-PAGE gels and subsequent in-gel trypsin digestion and MALDI-TOF MS. The protein was identified as peptidyl-prolyl isomerase B (PPI-B). In conclusion, the study demonstrates that SELDI-technique can be used not only to obtain islet protein patterns but is also helpful in the subsequent identification of differentially expressed proteins.

SELDI‐TOF as a method for biomarker discovery in the urine of aristolochic‐acid‐treated mice

“…An additional problem is the less-effective ionization in the high molecular range resulting with apparently lower sensitivity of the instrument for large proteins. After improvement of both the hardware and software, and thorough investigations of sample storage and preparation, SELDI-TOF MS is again being more frequently used [16][17][18][19]. The big advantage of the SELDI-TOF technology is the simple automation of sample preparation and the measurement process that enables its use for high-throughput screening, which is necessary when dealing with large sample numbers during the biomarker discovery process.…”

Proteomics in diabetes research

“…However, we have restricted ourselves not to include the field of diabetic complications, which are reviewed by others (Merchant and Klein 2007;Quin, et al 2007;Thongboonkerd and Malasit 2005). Neither have we included discussions on the techniques used for proteomic studies but the interested reader may look at references (Cho 2007;D'Hertog, et al 2006;Gorg, et al 2000;Keshamouni, et al 2006;Ortsäter, et al 2007). M a n u s c r i p t 6…”

Metabolites of Purine Nucleoside Phosphorylase (NP) in Serum Have the Potential to Delineate Pancreatic Adenocarcinoma

“…This has motivated multiple groups to profile the pancreatic juice proteome using both qualitative and quantitative mass spectrometry. These include Surface-Enhanced Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (SELDI TOF MS) [29], qualitative 2-Dimensional Electrophoresis (2DE)-based mass spectrometry [30] and quantitative Isotope-Coded Affinity Tags (ICAT)-labeled mass spectrometry [18]. Each of these has identified subsets of proteins that are altered in pancreatic cancer compared to non-cancer controls.…”