Using a 1 fg/mL baseline for limit of detection, single molecule FLISA demonstrated good qualitative agreement with gold standard tests on 19 of 20 patients, including accurately predicting the gold-standard-negative patients, while one gold-standard-positive patient produced no observable LAM in urine. Fluorescently labeled streptavidin revealed single molecule emission by epifluorescence microscope. Plasmonic gratings produced by low-cost soft lithography were bound with anti-LAM capture antibody, incubated with patient urine samples, and biotinylated detection antibody. GeneXpert MTB/RIF and culture testing of patient sputum confirmed the presence or absence of pulmonary tuberculosis while all patient urines were reference ELISA LAM-negative. METHODS AND FINDINGS:In this study, twenty sputum and urine sample sets were selected retrospectively from a repository of HIV-negative patient samples collected before initiation of anti-tuberculosis therapy. In this work, economical plasmonic gratings were used to analyze tuberculosis biomarker lipoarabinomannan (LAM) from clinical urine samples by single molecule fluorescence assay (FLISA) and compared with gold standard sputum GeneXpert MTB/ RIF, culture, and reference ELISA testing results. Finding methods to interrogate noninvasive, non-sputum clinical specimens is indispensable to improving access to tuberculosis diagnosis and care. Sputum is widely used as the testing input, but limited by its complexity, heterogeneity, and sourcing problems. The question whether the assay is suitable as a supplemental device in the diagnosis of HIV-associated TB, requires further investigations.BACKGROUND:Timely diagnosis of tuberculosis disease is critical for positive patient outcomes, yet potentially millions go undiagnosed or unreported each year. Conclusion: This commercially available generation of LAM-ELISA does not appear to be useful as an independent diagnostic test for pulmonary tuberculosis. Correlation with urinalysis revealed that proteinuria was significantly and positively associated with LAM-positivity (P = 0.026). The specificity amounted to 87.8% and was determined in participants with negative results in all microbiological tests and with sustained recovery under antibiotic treatment at day 56.
The sensitivity was noticeably higher in females (66.7%) and in HIV positive participants (62.0%). Results: Only 35 out of 69 pulmonary TB cases-confirmed by smear microscopy and/or solid culture and/or liquid culture-showed at least one positive LAM-ELISA result (sensitivity 50.7%). The participants were subsequently assigned to classification groups according to microbiological, clinical and radiological findings at recruitment and during a maximum follow up period of 56 days. Methods: The test was applied to two urine samples from 291 consecutively enrolled Tanzanian patients with suspected pulmonary tuberculosis.
In the present study, the now commercially available assay has been clinically assessed regarding its diagnostic value alone and in combination with clinical co-factors.
In a previous study, the first release of a urine LAM-ELISA by Chemogen (Portland, USA) has been evaluated with a promising sensitivity and specificity for the diagnosis of pulmonary TB. Background: The development and evaluation of rapid and accurate new diagnostic tools is essential to improve tuberculosis (TB) control in developing countries.
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