Work flow illustrating the steps involved in the proteomic analysis of OA synovial fluid. OA synovial fluid samples were pooled and subjected to depletion of abundant proteins by MARS-6 LC column and lectin affinity chromatography using three different lectins (Concanavalin A, wheat germ agglutinin and jacalin). The depleted fraction was then subjected to SDS-PAGE, SCX and OFFGEL fractionation. The lectin enriched fraction was subjected to SDS-PAGE analysis and SCX fractionation. All fractions were analyzed on LTQ-Orbitrap Velos mass spectrometer. Sequest and Mascot algorithms were used to perform database searches. Subsequently, gene ontology-based functional characterization of the identified synovial fluid proteins was carried out. Further, validation by MRM- based assays was carried out for three proteins identified from discovery studies.