Electrochemical biosensor for quantitation of anti-DNA autoantibodies in human serum

Abstract

Measurement of serum autoantibody is a critical tool in the diagnosis and management of autoimmune diseases. However, rapid and convenient methods at the point-of care have not been achieved in large part because any one antibody species is a heterogeneous and miniscule fraction of the total serum immunoglobulin displaying identical properties other than its antigen-binding specificity. The present system addresses these challenges by vacuum-mediated transport of diluted serum through an antigen-coated porous membrane. To measure anti-DNA autoantibodies, native DNA was immobilized into a poly(vinylidene fluoride) membrane pre-coated with a synthetic phenylalanine/lysine co-polymer. Flow-through of primary and peroxidase-conjugated secondary antibodies over the course of 3min enhanced productive antibody–antigen interactions by bringing the reactants into close mutual proximity. Signal was quantified electrochemically during the enzymatic conversion of the tetramethylbenzidine substrate to a charge-transfer complex. The electrochemical signals generated by sera from patients with systemic lupus erythematosus using this device showed good quantitative correlation with a standard enzyme-linked immunosorbent assay and displayed similar detection limits. Inter- and intra-assay variability and electrode uniformity were favorable as was a two-month test of the stability of the DNA-coated membrane. While refining the fluidics requirements of this biosensor will be needed, its capacity to quantify over the course of 30min anti-DNA antibodies in fresh human serum without background reactivity of normal serum makes this a promising technology as a point-of care device of clinical utility.