The ability to detect one specific biomarker in a pool of related antigens is critical for the unambiguous diagnosis of a given disorder or its state. The detection is not limited to active pathogenic forms. For example, one third of the world's population have a latent infection of tuberculosis, according to the World Health Organization (WHO). This emphasises the need for differential approaches which will allow systemic identification of a specific entity among multiple alternatives.
Directed biomolecular immobilisation
Reliability and quantitative reproducibility remain among the main challenges that prevent the routine use of differential diagnostics and hinder the development of IVD (in vitro diagnostic) and POC (point of care) devices.
Most tests are optimised as surface-based, which ensures straightforward readout and conversion of affinity constants into measurement values. One generic problem is the loss of activity of immobilised biomolecules due to their uncontrolled and multifocal attachments. This is characteristic of all surface-based assays that continue to suffer from batch-to-batch variations, proving costly for clinics.
NPL is developing a measurement methodology to enable quantitation and prediction of biological responses as a function of directed or controlled biomolecular immobilisation. In a recent study we have shown that detection of the interferon gamma (IFNγ) - a measure of the progression of diseases including tuberculosis - can be drastically enhanced. Together with partners from industry and healthcare, we are developing a new approach using model surfaces that immobilise and control the orientation of the IFNγ antibodies. This enables quantitative IFNγ capture at very low (nanomolar) concentrations in complex biological matrices such as cell culture and serum.
The validation of new biomarkers is an expensive and difficult process, which requires the comparison of analyte levels among many individuals over time. When no unequivocal single biomarker can be found, measurement of more than one analyte may be helpful to assess the efficacy of a given therapy or to distinguish one medical condition from another.
NPL can offer quantitative multiplex assays, i.e. tests that are able to quantify several analytes simultaneously. Specifically, our assays use in-line or 'single test line' multiplexing which allow for an increased number of individual assays per test. The assays can help validate new biomarkers and biochemical profiles as well as point-of-care tests.
One example is the monitoring of the wound status in diabetic foot ulcers using RegeniTherix™ - a proprietary 'smart dressing' technology developed by Neotherix. This study uses fluorescent lateral flow assays that utilise portable readers for quantitative determination of multiple analytes (inflammation markers IL6 and TNFa), which is in contrast to conventional lateral flow tests that tend to rely on visual interpretation of the test line's colour and thresholding.
Optical scattering artifacts observed in the development of multiplexed surface enhanced Raman spectroscopy nanotag immunoassays
Noble J, Attree S, Horgan A, Knight A, Kumarswami N, Porter R, Worsley G
Anal. Chem., 2012, 84, 8246-8252.
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