Prior to the western blotting experiment protein concentration in each sample [malaria patients (n = 24), FC (n = 6) and HC (n = 12)] was accurately estimated using the 2D-Quant kit (GE Healthcare) and BCA Protein Assay (Thermo Fisher Scientific). Western blot analysis was performed as described previously [40] . Briefly, serum proteins were separated by 12% SDS-PAGE (50 mg per track) and then transferred onto PVDF membranes under semidry conditions by using ECL semi-dry transfer unit (GE Healthcare). Western blot was performed by using monoclonal/ polyclonal antibody against serum amyloid A (Santacruz Biotechnology, sc-20651), haptoglobin (Santacruz Biotechnology, sc-71207), clusterin (Santacruz biotechnology, sc-8354) and retinolbinding protein (RBP) (Santacruz Biotechnology, sc-69795) and appropriate secondary antibody conjugated with HRP (GeNei (MERCK)-621140380011730 or 621140680011730). Candidate proteins for validation were selected on the basis of fold changes, possible association of the proteins with malaria pathobiology and accessibility of the required antibodies. ImageQuant software; version 5.0 (GE Healthcare) was applied for quantitation of signal intensity of the bands in western blots.