RESUMEN
Peptidomics research is of great significance for discovering potential biomarkers and monitoring human diseases. As a kind of common clinical biofluid, saliva known for its noninvasive collection and easy accessibility has been widely used in peptidomics research. In this article, we combined immobilized metal ions affinity chromotography (IMAC) with mesoporous material and proposed the copper ion doped magnetic mesoporous silica material (denoted as Fe3O4@mSiO2-Cu2+) which had a large surface area of 221â¯m2â¯g-1 and pore volume of 0.20â¯cm3â¯g-1. By immobilizing copper ions onto the mesopore walls, the standard peptide Angiotensin II could be identified in an extremely low concentration of 0.1â¯fmol⯵l-1 and in a mass ratio of 1:500 (Angiotensin II:BSA, m/m), which indicated significant sensitivity and a great size-exclusive ability. In addition, the introduction of polydopamine (PDA) made Fe3O4@mSiO2-Cu2+ more hydrophilic and biocompatible which could improve the profiling of endogenous peptides in bio-sample. Finally, 131 endogenous peptides were identified in human saliva after enrichment with Fe3O4@mSiO2-Cu2+. Therefore, Fe3O4@mSiO2-Cu2+ nanoparticles provided a promising candidate protocol for biomarker discovery.
Asunto(s)
Cromatografía Liquida , Cobre/química , Nanopartículas de Magnetita/química , Péptidos/análisis , Saliva/química , Dióxido de Silicio/química , Espectrometría de Masas en Tándem , Angiotensina II/química , Animales , Bovinos , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Péptidos/química , Porosidad , Albúmina Sérica Bovina/químicaRESUMEN
Variations in salivary components are closely associated with the predisposition and state of disease, the abnormal changes of salivary glycopeptidome are usually discovered as perilous singals of serious disease. Therefore, the monitoring and analyzing of salivary glycopeptidome are of even more overriding importance. In this work, a low-cost layer-by-layer assembly strategy was adopted to fabricate a hydrophilic multilayer magnetic probe (dubbed Mag-m-G6P) for salivary glycopeptidome analysis. The successful construction of multilayer structure not only guaranteed the good dispersal of probe by protecting magnetic core from itself aggregation tendency, but also endowed the probe with multiple advantages including the good hydrophilicity, uniform mesopore size and strong magnetic responsiveness, etc. As expected, with the optimized experimental conditions, the multifunctional probe showed high enrichemnt sensitivity, unbiased enrichment ability, excellent size-exclusion ability and reusability and so on in the process of standard sample analysis. At last, the Mag-m-G6P was successfully applied to salivary glycopeptidome analysis on further combination with LC-MS/MS analysis, a total of 53 endogenous glycopeptides were identified from human saliva.
Asunto(s)
Biología Computacional/métodos , Glicopéptidos/análisis , Magnetismo , Saliva/química , Cromatografía Liquida , Glicopéptidos/química , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Espectrometría de Masas en TándemRESUMEN
As one of the most important post-translational modifications (PTMs) of peptidome, glycopeptidome is closely related to serious disease, especially to cancer. In order to specifically discover and analyze glycopeptidome biomarkers for clinical diagnosis of cancer on early-stage, it is crucial to develop efficient technique to analyze low-abundance of endogenous glycopeptides in biological samples. In this report, a hydrophilic probe in mesoporous pore (denoted as Fe3O4@mSiO2@G6P) was designed and prepared. By taking advantage of the excellent hydrophilicity and size-exclusion ability, we applied Fe3O4@mSiO2@G6P to capture glycopeptides from both horseradish peroxidase (HRP) and immunoglobulin (IgG) digests successfully. Moreover, a total of 39 and 25 endogenous glycopeptides were identified from healthy saliva and gastric saliva, respectively, indicating the great potential of this probe for the exploration of glycopeptidome biomarkers.