title: An assessment of the indirect high intensity ultrasonic assisted cleavage of complex proteomes with immobilized trypsin.
abstract: The indirect high intensity ultrasonic assisted cleavage of complex proteomes using immobilized trypsin has been assessed. It has been found that the formation of aggregates between the beds supporting the immobilized trypsin is promoted. This affects the efficiency of the digestion process, which can be 100 times lower than the digestion efficiency obtained with in-solution trypsin. Through the use of isotopic peptide labelling with 18-O, it has been quantified that the digestion efficiency over serum samples can be 1.2-100 times higher for the 70% of the peptides when indirect ultrasonication is replaced by direct ultrasonication. Therefore, direct high intensity ultrasonic assisted cleavage of proteins is proposed as an alternative to be combined with immobilized trypsin.
authors.affiliation: Rheumatology Division, Proteomics Group-ProteoRed/ISCIII, INIBIC-CHUAC, A Coruña, Spain.
authors.forename: C
authors.initials: C
authors.lastname: Fernández-Costa
authors.forename: C
authors.initials: C
authors.lastname: Ruiz-Romero
authors.forename: F J
authors.initials: FJ
authors.lastname: Blanco
authors.forename: H M
authors.initials: HM
authors.lastname: Santos
authors.forename: J L
authors.initials: JL
authors.lastname: Capelo
mesh_terms: D016022:Case-Control Studies; D004800:Enzymes, Immobilized / Q000737:chemistry*; D006801:Humans; D007553:Isotope Labeling; D010003:Osteoarthritis / Q000097:blood*; D010103:Oxygen Isotopes; D010446:Peptide Fragments / Q000097:blood*; D059748:Proteolysis / Q000528:radiation effects; D020543:Proteome / Q000032:analysis*; D013010:Sonication / Q000379:methods*; D013016:Sound; D014357:Trypsin / Q000737:chemistry*
chemical_list: D004800:Enzymes, Immobilized; D010103:Oxygen Isotopes; D010446:Peptide Fragments; D020543:Proteome; D014357:Trypsin