Analyte multiplexing : Research : Biotechnology : Science + Technology : National Physical Laboratory

Analyte multiplexing

Single biomarkers for identifying medical conditions and for measuring the effectiveness of therapies are relatively rare. The validation of new ones is an expensive and difficult process requiring comparison of analyte levels among many individuals over time. When no unequivocal single biomarker can be found because values among individuals are too variable, measurement of more than one analyte may be helpful to confirm a therapy is working or distinguish one medical condition from another. In such cases the relative amounts of each analyte may be as important as the absolute values of the analytes. As researchers manage to find and validate new biomarkers and biochemical profiles, point-of-care tests that are able to quantify several analytes simultaneously (i.e. multiplex) in a quantitative manner will become increasingly valuable.

By adapting the widely used lateral flow test (e.g. Clearblue^® preganancy test) to use quantum dots, Raman nanotags (see figure) and fluorescent microspheres as spatially or spectrally addressable labels, the Biotechnology Group has been able to quantify multiple analytes within a single inexpensive disposable test. In contrast to conventional lateral flow tests, which tend to rely on visual interpretation of the test line's colour and thresholding, the NPL assays utilise handheld readers for quantitative determination of each analyte.

Scanning electron microscopy (SEM) image of surface enhanced Raman scattering (SERS) nanoparticles within the test line of a lateral flow immunoassay strip. The Raman spectra of the SERS nanoparticles depend on the dye that is adsorbed to each nanoparticle's gold core. Covalent immobilisation of antibodies onto the silica shells of the SERS nanoparticles allows analyte-specific binding. Positive signals are detected at the test line when an appropriate antibody adsorbed to the nitrocellulose substrate at the test line binds to its conjugate (analyte antibody-labelled SERS nanoparticle) to form an immuno-sandwich. Different analytes can be detected at the same test line and their amounts determined through deconvolution of the combined SERS spectra.