Research Paper Volume 12, Issue 23 pp 23698—23726

Figure 5. Proteomics workflow. An animal model of PS-induced diabetes-associated cognitive decline was established by exposure to three chronic psychological stressors. Fresh brain tissue MAM was extracted using gradient centrifugation, sample lysis was performed, and proteins were detected by an iTRAQ 8-plex. Following digestion and labelling, the samples were pooled, the peptides were fractionated by EASY-nLC1000 chromatography, and fractions were subsequently analyzed by standard LC-MS/MS. Peptides were identified and quantified based on their iTRAQ reporter area and relative protein quantification was inferred from these values. Key proteins were screened based on bioinformatics analysis and public databases were searched to further screen for differentially expressed proteins closely related to disease, for validation.