Ways to Reach Lower Detection Limits of Lateral Flow Immunoassays

Abstract

This chapter considers factors influencing sensitivity of lateral flow immunoassay and modern developments that are focused on reaching lower detection limits. The existing variety of proposed approaches is classified in accordance with the "big five rules" for these assays, including proper sample, receptor, interaction, response, and output. The solutions for rapid extraction of target analytes and preventing negative influence of extractants are considered. Role to antibodies affinity and specificity is characterized. Potential of alternate bioreceptor molecules is discussed. Immunoreactants' compositions, concentrations, and locations on the test strip are characterized as factors determining assay parameters. The existing variety of labels is compared in terms of their optical and alternate registration. Tools to modulate a sequence of analytical reactions and to form aggregates of the detected labels are considered. The discussed approaches are illustrated through developments of test strips for detection of mycotoxins, veterinary drugs, and other analytes.The overall design of the immunochromatographic test strip is shown in Figure 1. It is a composite of several membranes of different structures and porosities, fixed on a support. The bundling of the test strip can vary, so it makes sense to consider its design based on what analytical tasks are being performed on its different sites.A. Typically, the lower portion of the test strip contains a sample pad. It ensures the absorption of sample components, in which the presence of the target analyte is checked.B. The following is a section with immunoreagents that are washed out during the analysis and move upward along with the components of the sample. As a rule, a conjugate pad forms this zone. It contains a conjugate of antibodies against the target analyte with a nanodispersed label-particles of colored latex, colloidal gold, and so on.The next two sections are located on the main working membrane of the test strip. C.First, there is a zone along which the movement of the absorbed components of the sample and the washed immunoreagents continues. During this movement, immune reactions occur, and specific intermolecular complexes are formed. D.Next, a mixture of reacted and unreacted molecules enters the binding area with immobilized immunoreagents. Depending on whether the target analyte was present in the sample and in what amount, binding of labeled immune complexes occurs in certain areas (in the traditional case, with the formation of narrow colored lines). Usually, additional reagents are located here to control the functionality of the test system. E. The upper part of the test strip with the final pad, usually structurally similar to the sample pad, ensures the further movement of the reaction mixture under the action of capillary forces and the washing of unreacted components from the underlying areas. These processes allow the label's binding to be evaluated correctly.Membrane components of the test strip are fixed on a plastic support and partia...

Sensitivity enhancement in lateral flow assays: a systems perspective

“…Even in conventional LFA development, navigating trade-offs in system performance typically requires a long process of manual optimization over many experimental parameters, including those related to reagents and reaction conditions, materials, and test signals. 105,106 For example, in conventional LFA development, reagent optimization requires (at least) screening for high-affinity antibodies, selecting colorimetric labels, assessing the efficacy of attachment chemistry, cross-testing the capture and detection reagents, and evaluating the impact of blocking, spotting, drying, and/or rehydrating reagents in porous materials. Reaction condition optimization can include evaluating with sample matrices, and various buffers (for storage, reaction, wash/rinse, blocking, and/or running), surfactants, and other additives; testing the impact of various temperature, salt, and pH conditions; and testing porous materials for specific and non-specific binding.…”

Design of Multiplex Lateral Flow Tests: A Case Study for Simultaneous Detection of Three Antibiotics

“…Due to the increased number of controlled compounds in medical diagnostics, food safety and ecological monitoring, there is a need for multiplex test systems [6,7]. The location of several binding lines with reactants of different specificity is the best technological solution for multiplex ICA [8][9][10][11]. Such tests for 2-8 analytes are widely used to determine antibiotics in milk, mycotoxins in grain, etc.…”

Development of a Lateral Flow Highway: Ultra-Rapid Multitracking Immunosensor for Cardiac Markers

“…The devices for these assays (test strips) typically comprise multimembrane composites with preapplied reagents. Contact of the test strip with the sample leads to the movement of the sample solution along the membranes, and to the binding of a detectable label in certain zones of the strip [5]. The duration of the analysis is determined by the time it takes for the solution to move and for specific labeled complexes to form in sufficient quantities.…”