Dataset Information

Rapid detection of Mycobacterium tuberculosis biomarkers in a sandwich immunoassay format using a waveguide-based optical biosensor.

ABSTRACT: Early diagnosis of active tuberculosis (TB) remains an elusive challenge, especially in individuals with disseminated TB and HIV co-infection. Recent studies have shown a promise for the direct detection of pathogen-specific biomarkers such as lipoarabinomannan (LAM) for the diagnosis of TB in HIV-positive individuals. Currently, traditional immunoassay platforms that suffer from poor sensitivity and high non-specific interactions are used for the detection of such biomarkers. In this manuscript, we demonstrate the development of sandwich immunoassays for the direct detection of three TB-specific biomarkers, namely LAM, early secretory antigenic target 6 (ESAT6) and antigen 85 complex (Ag85), using a waveguide-based optical biosensor platform. Combining detection within the evanescent field of a planar optical waveguide with functional surfaces that reduce non-specific interactions allows for the ultra-sensitive and quantitative detection of biomarkers (an order of magnitude enhanced sensitivity, as compared to plate-based ELISA) in complex patient samples (urine, serum) within a short time. We also demonstrate the detection of LAM in urine from a small sample of subjects being treated for TB using this approach with excellent sensitivity and 100% corroboration with disease status. These results suggest that pathogen-specific biomarkers can be applied for the rapid and effective diagnosis of disease. It is likely that detection of a combination of biomarkers offers greater reliability of diagnosis, rather than detection of any single pathogen biomarker. NCT00341601.

SUBMITTER: Mukundan H

PROVIDER: S-EPMC3432392 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Rapid detection of Mycobacterium tuberculosis biomarkers in a sandwich immunoassay format using a waveguide-based optical biosensor.

Mukundan Harshini H Kumar Sandeep S Price Dominique N DN Ray Sonja M SM Lee Ye-Jin YJ Min Seonyeong S Eum Seokyong S Kubicek-Sutherland Jessica J Resnick Jesse M JM Grace W Kevin WK Anderson Aaron S AS Hwang Soo Hee SH Cho Sang Nae SN Via Laura E LE Barry Clifton C Sakamuri Ramamurthy R Swanson Basil I BI

Tuberculosis (Edinburgh, Scotland) 20120617 5

Early diagnosis of active tuberculosis (TB) remains an elusive challenge, especially in individuals with disseminated TB and HIV co-infection. Recent studies have shown a promise for the direct detection of pathogen-specific biomarkers such as lipoarabinomannan (LAM) for the diagnosis of TB in HIV-positive individuals. Currently, traditional immunoassay platforms that suffer from poor sensitivity and high non-specific interactions are used for the detection of such biomarkers. In this manuscript ...[more]

PMID: 22710249

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Project description:Improved access to effective tests for diagnosing tuberculosis (TB) has been designated a public health priority by the World Health Organisation. In high burden TB countries nucleic acid based TB tests have been restricted to centralised laboratories and specialised research settings. Requirements such as a constant electrical supply, air conditioning and skilled, computer literate operators prevent implementation of such tests in many settings. Isothermal DNA amplification technologies permit the use of simpler, less energy intensive detection platforms more suited to low resource settings that allow the accurate diagnosis of a disease within a short timeframe. Recombinase Polymerase Amplification (RPA) is a rapid, low temperature isothermal DNA amplification reaction. We report here RPA-based detection of Mycobacterium tuberculosis complex (MTC) DNA in <20 minutes at 39 °C. Assays for two MTC specific targets were investigated, IS6110 and IS1081. When testing purified MTC genomic DNA, limits of detection of 6.25 fg (IS6110) and 20 fg (IS1081)were consistently achieved. When testing a convenience sample of pulmonary specimens from suspected TB patients, RPA demonstrated superior accuracy to indirect fluorescence microscopy. Compared to culture, sensitivities for the IS1081 RPA and microscopy were 91.4% (95%CI: 85, 97.9) and 86.1% (95%CI: 78.1, 94.1) respectively (n = 71). Specificities were 100% and 88.6% (95% CI: 80.8, 96.1) respectively. For the IS6110 RPA and microscopy sensitivities of 87.5% (95%CI: 81.7, 93.2) and 70.8% (95%CI: 62.9, 78.7) were obtained (n = 90). Specificities were 95.4 (95% CI: 92.3,98.1) and 88% (95% CI: 83.6, 92.4) respectively. The superior specificity of RPA for detecting tuberculosis was due to the reduced ability of fluorescence microscopy to distinguish Mtb complex from other acid fast bacteria. The rapid nature of the RPA assay and its low energy requirement compared to other amplification technologies suggest RPA-based TB assays could be of use for integration into a point-of-care test for use in resource constrained settings.

Dataset Information

Rapid detection of Mycobacterium tuberculosis biomarkers in a sandwich immunoassay format using a waveguide-based optical biosensor.

Publications

Rapid detection of Mycobacterium tuberculosis biomarkers in a sandwich immunoassay format using a waveguide-based optical biosensor.

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