tailieunhanh - Báo cáo y học: "Immobilized pH gradient-driven paper-based IEF: a new method for fractionating complex peptide mixtures before MS analysis"

Tuyển tập các báo cáo nghiên cứu về y học được đăng trên tạp chí y học Critical Care giúp cho các bạn có thêm kiến thức về ngành y học đề tài: Immobilized pH gradient-driven paper-based IEF: a new method for fractionating complex peptide mixtures before MS analysis. | Seshi et al. Clinical Proteomics 2011 8 10 http content 8 1 10 CLINICAL PROTEOMICS RESEARCH Open Access Immobilized pH gradient-driven paper-based IEF a new method for fractionating complex peptide mixtures before MS analysis Beerelli Seshi1 Kumaraguru Raja1 2 and KH Chandramouli1 3 Correspondence BSeshi@ department of Pathology and Laboratory Medicine Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center 1124 West Carson Street Torrance California 90502 USA Full list of author information is available at the end of the article 2 BioMed Central Abstract Introduction The vast difference in the abundance of different proteins in biological samples limits the determination of the complete proteome of a cell type requiring fractionation of proteins and peptides before MS analysis. Methods We present a method consisting of electrophoresis of complex mixtures of peptides using a strip of filter paper cut into 20 sections laid end to end over a 24-cm-long IPG strip the pH gradient of which would drive the electrophoresis. Peptides absorbed onto individual paper pads after electrophoresis are subsequently recovered into a buffer solution thus dividing a complex peptide mixture according to pI into 20 liquid fractions. This paper-based IEF method PIEF was compared side-by-side with a similar but liquid-based Offgel electrophoresis OGE by analyzing iTRAQ-labeled peptide mixtures of membrane proteins from four different cell types. Results PIEF outperformed OGE in resolving acidic peptides whereas OGE did a better job in recovering relatively basic peptides. OGE and PIEF were quite comparable in their coverage identifying almost equal number of distinct proteins PIEF 1174 OGE 1080 . Interestingly however only 675 were identified by both of them each method identifying many unique proteins PIEF 499 OGE 415 . Thus the two methods uncovered almost 40 more proteins compared to what is obtained by only one method. .