Microfluidic barcode assay for antibody-based confirmatory diagnostics.
Confirmatory diagnostics offer high clinical sensitivity and specificity typically by assaying multiple disease biomarkers. Employed in clinical laboratory settings, such assays confirm a positive screening diagnostic result. These important multiplexed confirmatory assays require hours to complete. To address this performance gap, we introduce a simple 'single inlet, single outlet' microchannel architecture with multiplexed analyte detection capability. A streptavidin-functionalized, channel-filling polyacrylamide gel in a straight glass microchannel operates as a 3D scaffold for a purely electrophoretic yet heterogeneous immunoassay. Biotin and biotinylated capture reagents are patterned in discrete regions along the axis of the microchannel resulting in a barcode-like pattern of reagents and spacers. To characterize barcode fabrication, an empirical study of patterning behaviour was conducted across a range of electromigration and binding reaction timescales. We apply the heterogeneous barcode immunoassay to detection of human antibodies against hepatitis C virus and human immunodeficiency virus antigens. Serum was electrophoresed through the barcode patterned gel, allowing capture of antibody targets. We assess assay performance across a range of Damkohler numbers. Compared to clinical immunoblots that require 4-10 h long sample incubation steps with concomitant 8-20 h total assay durations; directed electromigration and reaction in the microfluidic barcode assay leads to a 10 min sample incubation step and a 30 min total assay duration. Further, the barcode assay reports clinically relevant sensitivity (25 ng ml(-1) in 2% human sera) comparable to standard HCV confirmatory diagnostics. Given the low voltage, low power and automated operation, we see the streamlined microfluidic barcode assay as a step towards rapid confirmatory diagnostics for a low-resource clinical laboratory setting.
Department of Bioengineering, University of California, Berkeley, Berkeley, CA, USA.
M Kursad
MK
Araz
Akwasi A
AA
Apori
Cleo M
CM
Salisbury
Amy E
AE
Herr
D000906:Antibodies / Q000276:immunology*; D019937:Diagnostic Techniques and Procedures; D004553:Electric Conductivity; D006801:Humans; D055767:Immobilized Proteins / Q000032:analysis / Q000737:chemistry; D007118:Immunoassay / Q000379:methods*; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D009068:Movement; D019809:Streptavidin / Q000378:metabolism; D013997:Time Factors
D000906:Antibodies; D055767:Immobilized Proteins; D019809:Streptavidin