Itraq 8 plex manual

 · In iTRAQ it was shown that, due to an increase in the average ion charge state, there is a significant reduction in the number of identifications. Further, the iTRAQ 8-plex was reported to result in reduced protein annotation rate in comparison to the iTRAQ 4-plex. During fragmentation, the loss of fragments of the label tag from precursor ions may occur, which causes some difficulties in the . Refer to the iTRAQ reagents 8-plex protocol for more details on the reagents (Page ). When instructed to inject a solution in the procedures: Fill a clean syringe (5 ml) with the indicated solution. Remove air bubbles. Insert the syringe needle (blunt tip) into the . Each vial of 8-plex reagent contains 2 µmol of iTRAQ reagent. This is usually sufficient to label µg of protein digest. For plasma and serum samples which contain many non-peptide primary amine compounds, the protocol may need to be adjusted.

Procedure. The ITRAQ method is based on the covalent labeling of the N-terminus and side chain amines of peptides from protein digestions with tags of varying mass. There are currently two mainly used reagents: 4-plex and 8-plex, which can be used to label all peptides from different samples/treatments. We employed UPLC-MS/MS with iTRAQ 8-plex labeling to quantitatively analyze the supernatant produced by two Chinese hamster ovary (CHO) cell lines (CHO K1SV and CHO CAT-S). In each case, the supernatant from the host and three transfected clones were analyzed at days 5, 7, and 10 of culture. A total of eight iTRAQ 8-plex experiments were performed. We used an 8-plex iTRAQ system to analyze the proteomes of hESCs during proliferation and at different stages of differentiation in three biological replicates. For each biological replicate, the experiments were repeated twice. Protein lysates from ESCs and EBs on days 6, 12, and 20 after initiation of differentiation were used for iTRAQ labeling.

Each vial of 8-plex reagent contains 2 µmol of iTRAQ reagent. This is usually sufficient to label µg of protein digest. For plasma and serum samples which contain many non-peptide primary amine compounds, the protocol may need to be adjusted. DRAFT am, 7x9_MultiChapter_www.doorway.ru iv iTRAQ™ Reagents Chemistry Reference Guide Chapter 4 Analyzing with a Q TRAP iTRAQ ™ Reagents. Contains iTRAQ ® Reagents , , , , , , and and sufficient material for five 8-plex assays. Multiplex Buffer kit (Cat. No. ) to be.