Positive and negative controls were run in duplicate. The plates were read immediately at 450 nm on an ELISA plate reader (Anthos Labtec HT3, Wals/Salzburg, Austria). The induced sputum supernatants were thawed and processed using the TB LAM ELISA kit according to the manufacturer’s instructions. Two × 1.5 ml aliquots of supernatant were frozen at −20☌ prior to batched LAM ELISA testing. It was then heated at 95☌ for 30 min, and centrifuged at 12 500 rpm for 30 min. An aliquot was removed for smear microscopy and liquid culture before filtering the sample through 2-ply sterile 50 μm gauze and centrifugal washing with phosphate buffered saline (PBS) at 1200 rpm 15 ml of the non-cellular fraction was collected. Induced sputum samples were liquefied by adding 2 × volume of 0.1% dithiothreitol and rolling for 20 min at room temperature to homogenise. 12 We therefore hypothesised that improving sputum quality by sampling the lower respiratory tract using induced sputum after mouth rinsing could potentially reduce oropharyngeal contamination, thereby improving assay specificity and offering potential clinical utility for the diagnostically challenging sputum-scarce patient with suspected TB.Īll induced sputum samples were processed within 2 h of collection. We demonstrated that the poor test specificity was likely due to cross-reactivity with LAM or LAM-like molecules found in oral mouth flora of TB suspects. We have recently studied the utility of the TB LAM ELISA using expectorated sputum and found good sensitivity (86%) but poor specificity (15%). The performance of the TB LAM ELISA has been evaluated in a number of non-urine biological samples such as serum samples, 8, 9 pleural fluid 10 and cerebrospinal fluid, 11 and found to offer limited clinical utility. 6 It has been shown to be a useful rule-in diagnostic test for TB using urine from HIV-infected patients with advanced immunosuppression. Its low cost and recent availability as a point-of-care lateral flow strip test make it particularly appealing for use in high-burden, resource-limited settings. 5 A commercially available LAM enzyme-linked immunosorbent assay (ELISA TB LAM ELISA, Alere, Waltham, MA, USA) was designed for the detection of LAM using urine as the target biological sample. 4 It is a heat-stable antigen released from active and degrading Mycobacterium tuberculosis. ![]() Lipoarabinomannan (LAM) is a 17.5 kDa lipopolysaccharide and immunogenic mycobacterial cell wall antigen. ![]() 3 There is an urgent need, therefore, particularly among HIV-co-infected patients, for a rapid and less expensive TB test suited to resource-poor settings. 2 Newer molecular diagnostic technologies, such as the Xpert ® MTB/RIF assay (Cepheid, Sunnyvale, CA, USA), appear promising, but are untested in sputum-scarce individuals and remain expensive and unavailable in most high-burden settings. ![]() ![]() Available TB diagnostic tools perform suboptimally in high HIV prevalent settings, with sensitivity of smear microscopy as low as 20%. Clinical disease presentation and radiological manifestations can be atypical, particularly in HIV-co-infected patients. 1 TB diagnosis is challenging, and current diagnostic tools are inadequate. TUBERCULOSIS (TB) kills 1.8 million people annually and is the leading cause of human immunodeficiency virus (HIV) related deaths in sub-Saharan Africa.
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