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

• Anti-DNA antibodies in human sera are an important diagnostic indicator of systemic lupus erythematosus.
• Serum anti-DNA antibodies and peroxidase-conjugated anti-IgG were transported through a permeable membrane to which DNA antigen was indirectly immobilized.
• We quantified IgG antibody activity in a total assay time of 30 min and propose its electrochemical basis.
• The results agreed well with those of a standard ELISA that required 5 h.
• Performance assessments regarding assay reproducibility, electrode uniformity, and storage of DNA-coated membrane were favorable.

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 3 min 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 30 min 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.