Glucose is the predominant source of energy in cells. However, a chronic high glucose exposure of proteins modifies a number of biological pathways, known as glucotoxicity. Several studies have suggested that this impaired protein function is associated in part to protein glycation. However, despite the evidence of this glucotoxicity on tissues and cells, the exact mechanisms underlying the loss of protein function by glycation are not well understood. Strategies that will allow the discovery of the identity and function of the glycated plasma proteins generated by chronic hyperglycemia would be of a considerable help to further understand the underlying mechanisms implicated in protein dysfunction associated with glucotoxicity. The present chapter describes an innovative and interdisciplinary proteomics strategy to achieve a comprehensive identification and quantification of glycated proteins in plasma. This should provide new molecular insights into protein glycation mechanisms and identify new targets to improve the subsequent defective protein function.