Proteomic analysis of serum from Chikungunya patients

Serum from Chikungunya patients was pooled and subjected to depletion using multiple affinity removal system to remove the abundant proteins. The depleted protein was then fractionated using SCX. All the fractions were analyzed on the high-resolution LTQ-Orbitrap Velos mass spectrometer. The data obtained was searched against the human RefSeq 52 proteins using Proteome Discoverer, version 1.3.0.339 workflow. The workflow consisted of spectrum selector and reporter ion quantification nodes in addition to SEQUEST and Mascot search nodes. Similar parameters were used in all the searches with trypsin as enzyme allowing a single missed cleavage. Other parameters include methylthiol modification of cysteine, iTRAQ labels at the peptide N-terminus and Lysine residues as static modifications and oxidation of methionine as variable modification. A mass tolerance of 20 ppm and 0.1 Da were used for the precursor ion and fragment ions, respectively with a signal to noise ratio of 1.5 for a precursor mass range of 350–10000 Da. A false discovery rate (FDR) of 1% was applied to the results.