Luminex assays are bead-based immunoassays that utilize principles similar to traditional sandwich ELISA assays. Luminex-based assays have multiplexing abilities and can quantify up to 100 different analytes in a single assay volume. Hence, they are used widely in the drug development process. To give an overview of the drug development process, Luminex-based assays are advantageous at every stage of drug development. They can effectively quantify multiple analytes and can help researchers understand complex biological and disease processes.
The main advantage of Luminex platforms is custom Luminex assays. Researchers can develop custom Luminex assays by tailoring assay components to their research needs. Many assay services have a pre-developed menu of analytes. However, researchers can test newer analytes or sample types through customized Luminex assays. Custom Luminex assays can help translate multiplex ideas into bioanalytical realities. However, with any custom development, there are specific assay features. The current article discusses aspects of custom Luminex-based assays for beginners.
Custom Luminex-based assay
Custom assay focuses on high performance, with each assay lot tested for consistent standard curves, low background, and wide dynamic range. Besides, most assay developers anchor each standard sample to the same master calibrator so that researchers can compare sample data over time.
A minimum detectable dose is the lowest value statistically different from zero. This value is critical in assessing assay sensitivity. Researchers calculate the minimum detectable dose by adding values of two standard deviations and the mean optical density of numerous zero standard replicates and then measuring the corresponding analyte concentration from the standard curve. Highly sensitive custom assays have a lower standard curve range. Assay developers design custom Luminex assays with low background, high signal, and enhanced sensitivity.
Sometimes assays may give false-positive results. False-positive means incorrectly detecting an analyte in study samples. Custom assays have linearity experiments to identify false-positive signals. Sample values must ideally remain consistent when running back-calculating concentration and multiple dilutions. Issues related to specificity may arise when sample values increase with rising dilutions. Custom assays are specifically built to prevent any false-positive results.
Besides, assay precision is another vital factor for any bioanalytical assay. Precision is defined as an assay’s ability to reproduce results between and within assays. Precision is critical to assure that an assay can provide consistent results among separate runs and thus find similarities or differences between two assay results. Inter-assay and intra-assay precision are two types of precision considered in bioanalytical assays. Intra assay assesses reproducibility within different wells in the same assay. Researchers run replicates to assess intra-assay reproducibility. Whereas inter-assay precision is reproducibility between separate assay runs. It helps confirm an assay’s ability to reproduce results using multiple kits over time. Ideally, developers achieve a CV value of less than 30% for most custom Luminex assays. Some high-performance assays also have a CV value of less than 17%. Such robust assays instill the required confidence in research.
Custom Luminex assays are ideal for analyzing novel analytes in biological samples. However, researchers must consider individual requirements and plan on developing a custom Luminex assay accordingly.