Detection of Lipoarabinomannan (LAM) antigen from the urine of tuberculosis-infected children by using Ceres Nanotrap® magnetic particles.
K. Ahmed1, T. Sattar1, N. T. Iqbal1,2 , T. Thomas3
1 Department of Pediatrics and Child Health, Aga Khan University, Karachi, 2 Department of Biological and Biomedical Sciences, Aga Khan University, Karachi, 3 Department of Medicine, University of Virginia, USA
Background: One-quarter of the world’s population is infected with tuberculosis (TB). Each year ten million people develop active TB, of which one million are children. According to the WHO-2019 report, 36% of new TB cases remain unreported or undiagnosed, due to the limitations of recent diagnostic tools which are insensitive or have low diagnostic performance. Lipoarabinomannan (LAM) is a TB biomarker that can be detected in urine. LAM is a lipopolysaccharide antigen found in the mycobacterial cell wall and is released from metabolically active or degenerating bacterial cells. In this study, we aim to increase the sensitivity and diagnostic accuracy of the LAM test by using Ceres Nanotrap particles (Ceres Nanosciences, Manassas, VA, USA) to capture specifically LAM antigen in urine specimens from GeneXpert positive children and compare it with age-matched control.
Methodology: The urine processing protocol was first optimized to capture the LAM antigen by using magnetic Ceres nano trap particles. Urine samples were also spiked with ATCC TB LAM antigen derived from M. leprae as a positive control, which was kindly gifted by BEI Resources. Briefly, urine samples from the cohort of GeneXpert positive children (n=12) aged between 1-18 years and their age-matched controls (n=10) from the community were tested on Alere TB LAM strips. Neat (unsorted and non-spiked) urine samples without being processed with Ceres Nano trap particles were also tested on the Alere TB LAM strip to check any background or false result. Urine samples were sorted using Ceres nano trap particles to augment the sensitivity, after following the processing steps, samples were loaded on Alere TB LAM strips. The appearance of a visible purple/gray band in the patient window was considered as positive for the presence of Mycobacteria LAM antigen in the urine sample.
Results: A significant difference was observed when urine samples were sorted with magnetic Ceres nanotrap particles in GeneXpert positive cases. The LAM test was found positive in 6/12 (50%) in sorted urine sample compared with controls 1/10 (10%). Whereas, in unsorted (neat) urine samples, no significant difference was found between cases 1/12(8%) and controls 1/10(10%)
Conclusion: Urine TB LAM assay is an attractive diagnostic biomarker test to detect TB. The approach of concentrating LAM antigen using Ceres nano trap particles enhances the algorithm of TB diagnosis and to achieve high sensitivity enough to reach and assistance to all TB patients.
WHO global tuberculosis report 2019. https://www.who.int/teams/global-tuberculosis-programme/global-report-2019