Multiple-Analyte Fluoroimmunoassay Using an Integrated Optical Waveguide Sensor

Abstract

A silicon oxynitride integrated optical waveguide was used to evanescently excite fluorescence from a multianalyte sensor surface in a rapid, sandwich immunoassay format. Multiple analyte immunoassay (MAIA) results for two sets of three different analytes, one employing polyclonal and the other monoclonal capture antibodies, were compared with results for identical analytes performed in a single-analyte immunoassay (SAIA) format. The MAIA protocol was applied in both phosphate-buffered saline and simulated serum solutions. Point-to-point correlation values between the MAIA and SAIA results varied widely for the polyclonal antibodies (R2 = 0.42-0.98) and were acceptable for the monoclonal antibodies (R2 = 0.93-0.99). Differences in calculated receptor affinities were also evident with polyclonal antibodies, but not so with monoclonal antibodies. Polyclonal antibody capture layers tended to demonstrate departure from ideal receptor-ligand binding while monoclonal antibodies generally displayed monovalent binding. A third set of three antibodies, specific for three cardiac proteins routinely used to categorize myocardial infarction, were also evaluated with the two assay protocols. MAIA responses, over clinically significant ranges for creatin kinase MB, cardiac troponin I, and myoglobin agreed well with responses generated with SAIA protocols (R2 = 0.97-0.99).

A Duplexed Microsphere-Based Fluorescent Immunoassay

“…Furthermore, low analyte and labeled antibody concentrations are needed to minimize nonspecific effects and interferences between the individual immunoassays of multiplexed assays. Nonspecific binding to the solid surface, cross-reactivity, and aggregation of the individual tracer molecules can decrease the performance of multiplexed immunoanalyses (9,15,34). It appears that it is more difficult to meet these requirements for competitive than for noncompetitive immunoassays because the latter assays are generally more sensitive (35).…”

Virus detection using filament‐coupled antibodies

“…A number of schemes based on the sandwich immunoassay have been reported in the literature. For example, several recent applications are based upon changes in electrical properties as antigen binds to an immobilized probe (Balasubramanian et al, 2004;Baselt et al, 1996;Mubammad-Tahir and Alocilja, 2003;Ronkainen-Matsuno et al, 2002) or based on the interaction of evanescent light waves with surface-bound antibodies (Ligler et al, 2003;Plowman et al, 1999;Rowe et al, 1999). Many of these approaches are either labor intensive or require complex microfluidic devices and are not well suited for point-of-care clinical diagnostics or for field testing for biohazardous materials.…”

Rapid Multiplexed Immunoassay for Simultaneous Serodiagnosis of HIV-1 and Coinfections

“…1). Building on a patented design introduced by Herron and colleagues (6,12,13,17), the MBio lens design overcomes the fundamental challenge of reproducible light coupling to the waveguide. The lens design creates a diverging beam; modes mix down the length of the waveguide, eventually creating a spatially uniform illumination field along the axial length of the waveguide.…”