Development of a new parallelized, optical biosensor platform for label-free detection of autoimmunity-related antibodies

Abstract

Autoimmune diseases are characterized by the presence of autoantibodies in serum of affected patients. The heterogeneity of autoimmune relevant antigens creates a variety of different antibodies, which requires a simultaneous detection mode. For this reason, we developed a tool for parallelized, label-free, optical detection that accomplishes the characterization of multiple antigen-antibody interactions within a single measurement on a timescale of minutes. Using 11-aminoundecyltrimethoxysilane, we were able to immobilize proteinogenic antigens as well as an amino-functionalized cardiolipin on a glass surface. Assay conditions were optimized for serum measurements with a single spot antigen chip on a single spot 1-λ detection system. Minimized background signal allows a differentiation between patients and healthy controls with a good sensitivity and specificity. Applying polarized imaging reflectometric interference spectroscopy, we evaluated samples from three APS patients and three control subjects for this proof-of-principle and already obtained good results for β2-glycoprotein I and cardiolipin.

Diagnostic performance study of an antigen microarray for the detection of antiphospholipid antibodies in human serum

“…Antigeneity of the covalently bound protein was confirmed by binding of a monoclonal anti-β 2 -GPI. Similar measurements with a monoclonal anti-transferrin gave almost no signal and therefore verify the specificity of the system (described in [15]). A titration experiment with anti-β 2 -GPI (5.0-40.0 μg/mL) is additionally shown in Supplemental Material Figure 1.…”

“…As a solution, the whole IgG fraction was isolated with recombinant protein A/G coated sepharose spin columns prior to the measurement. described in Bleher et al [15]). Transducer chips for the pi-RIfS system are in the dimension of a standard microscope slide (75 × 25 mm).…”

“…pi-RIfS Our biosensor setup, as well as the measurement performance, was described in detail by Bleher et al [15]. A summary of the technical features, including data processing, is given in the Supplemental Material.…”

“…Since a proof of principle of the underlying detection method has already been published by our group in Bleher et al [15], we now focussed on the validation of the microarray to measure larger patient collectives. We further characterized the array in terms of sensitivity and specificity in comparison to standard APS antibody detection performed with ELISA systems.…”

Plattformtechnologie für die mobile, markerfreie Proteindetektion

“…Dafür sind kompakte, schnell arbeitende, dezentrale und einfach zu benutzende Sensoren von hohem Interesse für Point-of-Care Anwendungen. Diese Sensoren sollten in der Lage sein, spezifisch mehrere Proteine für eine eindeutige Diagnose nachzuweisen [3][4][5]. Im Gegensatz zu markerbasierten Messmethoden wie z.B.…”