Over the years, lateral flow assays (LFAs) have become an indispensable element of the rapid, point-of-care diagnostic test market. Initially gaining prominence through their early implementation in home pregnancy test kits, lateral flow immunoassays have since diversified to encompass a wide array of devices designed for point-of-care settings, removing the need to send samples to a laboratory or clinic for analysis. These tests skillfully detect an extensive range of biomarkers, pathogens, and environmental contaminants in numerous industries, making them an invaluable tool in modern diagnostics.
The underlying technology of lateral flow assays dates back over three decades, yet these systems have continually evolved to meet the complex diagnostic demands of today. State-of-the-art lateral flow assays boast high sensitivity and multiplexing capabilities, effectively addressing the high error rate that often arises at the point of sampling. By incorporating user-centric designs, LFAs have become more accessible and easier to use, even for individuals with minimal technical expertise.
The scope of these assays now encompasses a wide variety of diagnostic devices, ranging from simple, cost-effective, and lower complexity qualitative tests to intricate designs that facilitate the quantification of multiple biomarkers in real-world settings. These advanced devices eliminate the need for capital-intensive equipment and infrastructure, enabling rapid and accurate diagnostics in various field applications.
In recent years, the development of customizable readers, smartphone-based readers, and digital platforms has further enhanced the capabilities and applications of LFAs. These advancements have not only improved the accuracy and reliability of test results but also enabled the integration of diagnostic data with electronic health records and other digital systems. This seamless data integration facilitates better decision-making, improved patient care, and more efficient resource allocation in healthcare settings.
The expanding applications of LFIs have led to their adoption in diverse sectors beyond healthcare, such as food safety, agriculture, environmental monitoring, and veterinary medicine. The versatility and adaptability of lateral flow assays make them a powerful diagnostic tool, with the potential to revolutionize various industries by delivering rapid, accurate, and accessible testing solutions.
Lateral flow assays, a popular format for Rapid Test Devices or RDT’s, are immunosensors that rely on the use of low cost materials to achieve the flow of reactants. At their simplest, they are dipsticks with a sample application pad at one end, a reading zone where the signal is generated in the middle and an absorbent material at the other end. Tests may be incorporated into housings (cassettes) that support higher functionality, branding and user-centric design goals.
Lateral flow immunoassays may be qualitative (indicating the presence or absence of an analyte), semi-quantitative, or fully quantitative. Each type of assay offers distinct advantages, depending on the specific diagnostic requirements and desired outcomes.
Qualitative lateral flow immunoassays are designed to indicate the presence or absence of an analyte in a sample. These assays are often used for rapid, preliminary screening purposes, providing a simple yes or no answer regarding the existence of a target substance. Such tests are particularly useful in situations where a quick assessment is needed, such as detecting the presence of pathogens, allergens, or drugs of abuse.
Semi-quantitative lateral flow immunoassays offer an intermediate level of information, providing an estimation of the analyte’s concentration within the sample. While not as precise as fully quantitative assays, semi-quantitative tests can still offer valuable insights into the severity or progression of a condition or the effectiveness of a treatment. This type of assay is frequently employed in monitoring chronic diseases, evaluating immune responses, or assessing environmental contamination levels.
Fully quantitative lateral flow immunoassays deliver the most comprehensive results, accurately measuring the exact concentration of an analyte within a sample. Quantitative assays are essential in situations where precise measurements are required to make informed decisions regarding diagnosis, prognosis, or treatment. These tests are commonly used in the monitoring of hormone levels, the quantification of specific biomarkers, and the determination of viral loads, among other applications.
Lateral flow assays contains several important components:
These materials, each serving one or more purposes, overlap onto one another and are mounted on a backing card using a pressure sensitive adhesive. Figure 1 shows a typical configuration.
Figure 1: Typical strip configuration, including a sample pad, particle conjugate on the conjugate pad, nitrocellulose membrane, wick and backing card.
When a lateral flow assay is run, a sample is added to a Sample Application Pad. Here, the sample is treated to make it compatible with the rest of the test. The treated sample migrates through this region to the Conjugate Pad. Here, a particulate conjugate has been immobilized, typically a colloidal gold or a colored, fluorescent, or paramagnetic monodisperse latex particle or cellulose nanobead. This particle has been conjugated to one of the specific biological components of the assay, either antigen or antibody depending on the assay format.
The sample re-mobilizes the dried conjugate, and the analyte in the sample interacts with the conjugate as they both migrate into the next section of the strip. This zone, known as the Reaction Matrix is a porous membrane onto which the other specific biological components of the assay have been immobilized. These are typically proteins, either antibody or antigen that have been laid down in bands in specific areas of the membrane where they serve to capture the target and conjugate as they migrate up the strip. Excess reagents move past the capture lines and are entrapped in the Wick or absorbent pad. Results are interpreted on the Reaction Matrix as the presence or absence of lines and can be read either by eye or using a reader.
Assay formats are either sandwich (direct) or competitive (competitive inhibition) in nature, and the assay format can accommodate qualitative, semi-quantitative, or fully quantitative assays. Direct assays are typically used when testing for larger analytes with multiple antigenic sites, such as hCG, Dengue antibody or antigen or HIV. In this case, a positive result is indicated by the presence of a test line. Less than an excess of sample analyte is desired, so that some of the conjugated particles will not be captured at the capture line, and will continue to flow toward the second line of immobilized antibodies, the control line. This control line typically comprises a species-specific anti-immunoglobulin antibody specific for the conjugated antibody.
Competitive formats are typically used when testing for small molecules with single antigenic determinants, which cannot bind to two antibodies simultaneously. In this format, a positive result is indicated by the absence of a test line on the reaction matrix. A control line should still form, irrespective of the result on the test line. The two formats are illustrated schematically in Figures 2a and 2b below.
Figure 2a: Direct Solid Phase Immunoassay
Figure 2b: Competitive solid phase immunoassay
Lateral flow immunoassays are easy to use. The user simply needs to dip the sample pad into the liquid they want to test and then lay the test flat or apply the sample directly to the cassette. If the test is positive, a colored or fluorescent line will show up on the test along with a control line. If the test is negative, only the control line will be present. For many years, women with no medical training have successfully used this kind of test to detect pregnancy or ovulation at home.
More advanced lateral flow tests provide additional benefits, including higher sensitivity, multiplexing and the ability to detect multiple biomarkers from a single finger stick. These advanced tests are useful for in-the-field testing.
Lateral flow rapid tests provide results quickly. In many cases, the test and control lines show up almost immediately with definitive results available within a few minutes. This rapidity is important in medical settings where doctors have limited time available to spend with each patient, as well as in manufacturing process quality control checks in pharmaceutical companies, highly sensitive human performance testing for athletes, and bioprocessing and GMO testing in agriculture and food safety.
DCN Dx is the premier choice for designing and developing point-of-care rapid diagnostic assays with challenging specifications, such as:
Our cross-functional of assay development, engineering, manufacturing, and IVD clinical research professionals allows us to assist you in the development of your entire rapid diagnostic test from concept to commercialization.
Our staff has unparalleled experience in developing the highest performing point-of-care assays in lateral flow markets, ranging from medical diagnostics to veterinary, food science, bio-defense, agricultural, and environmental health tests.
DCN Dx operates under ISO 9001:2015 and ISO 13485:2016. All of our development programs are done under Design Controls, and all production activities for reagents, components, assays, and electronics are performed under our EN 13485 system.