Immunoassay Development
Immunoassay Development - Related Videos
09:08
Developing a Salivary Antibody Multiplex Immunoassay to Measure Human Exposure to Environmental Pathogens
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Cited by 11 •
2016
In the current climate of scarce resources, new technologies are emerging that allow researchers to conduct studies cheaper, faster and with more precision. Here we describe the development of a bead-based salivary antibody multiplex immunoassay to measure human exposure to multiple environmental pathogens simultaneously.
09:02
Fabricating a UV-Vis and Raman Spectroscopy Immunoassay Platform
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Cited by 4 •
2016
Nanoparticle-based optical probes have been designed as a vehicle for detecting antigens using Raman and UV-Vis spectroscopy. Here we describe a protocol for preparing such probes for a UV-Vis/Raman spectroscopy immunoassay in such a way to incorporate future multiplexing capabilities.
11:33
Fabrication of Three-dimensional Paper-based Microfluidic Devices for Immunoassays
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Cited by 12 •
2017
We detail a method to fabricate three-dimensional paper-based microfluidic devices for use in the development of immunoassays. Our approach to device assembly is a type of multilayer, additive manufacturing. We demonstrate a sandwich immunoassay to provide representative results for these types of paper-based devices.
08:05
Multiplexed Fluorometric ImmunoAssay Testing Methodology and Troubleshooting
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Cited by 19 •
2011
Using Luminex Corporation’s xMAP microsphere technology, we have developed the Multiplexed Fluorometric ImmunoAssay (MFIA) for serosurveillance of various laboratory animal species. The MFIA is a suspension microarray where antigen, tissue control or immunoglobulins are covalently linked to color-coded polystyrene microspheres. The MFIA testing method as well as various troubleshooting topics is addressed.
07:32
A Capillary Electrophoresis Immunoassay for Protein Biomarker Detection
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2025
This video demonstrates a capillary electrophoresis immunoassay protocol for detecting a protein biomarker associated with amyotrophic lateral sclerosis (ALS) from human blood platelets. Confirmation of the presence of the ALS biomarker protein is achieved through size-based separation of the platelet proteins followed by chemiluminescent detection.
FAQs
Antibody-pair selection strongly influences binding specificity, sensitivity, and signal quality. Developers compare antibodies for their ability to recognize the intended target and work together within the selected assay format. A suitable pair improves target measurement, whereas poorly matched antibodies can reduce signal performance or increase experimental variability. This decision is therefore central to producing reproducible analytical results.
Nonspecific binding produces signal that does not represent the intended target, making measurements less accurate and less reproducible. During optimization, developers adjust assay components and reaction conditions to reduce these unwanted interactions while preserving specific antigen–antibody binding. Controlling this balance improves specificity and helps distinguish genuine target-associated signal from background signal in research or testing.
Assay formats determine how target binding is organized and how the resulting signal is measured. Detection labels provide the signal used to identify or quantify the target, so their selection affects signal strength and interpretation. Developers evaluate these choices together with antibody pairs and reaction conditions to obtain a format that supports reliable qualitative or quantitative measurements.
Sensitivity indicates how effectively an assay detects or measures the target, while calibration links observed signal to the reported measurement. Signal performance determines whether that relationship is sufficiently consistent for interpretation. Evaluating all three helps developers judge whether an assay can provide dependable quantitative results rather than merely producing a detectable response.
A typical workflow begins by selecting antibody pairs and an assay format, followed by choosing detection labels and establishing reaction conditions. Developers then optimize binding and signal behavior, assess specificity and sensitivity, and evaluate calibration and reproducibility. Iterative testing is important because changes to one component can alter the assay’s overall measurement performance.
Developers examine reaction conditions that influence antigen–antibody binding, nonspecific interactions, and signal generation. They also assess how those conditions affect calibration, sensitivity, and reproducibility across measurements. Systematic optimization helps identify a stable combination of assay components and conditions, reducing experimental variability and supporting consistent results in later research, diagnostic, or quality-control use.
In bioengineering, optimized immunoassays support biomarker analysis, diagnostic device development, therapeutic monitoring, and characterization of engineered proteins or cells. Their value comes from converting specific molecular recognition into qualitative or quantitative data. Reliable assay performance can therefore help researchers evaluate biological samples, monitor engineered systems, and support bioprocess quality control.