Development of antigen detection assay for diagnosis of tuberculosis-usingsputum samples

The rising incidence of tuberculosis worldwide means an increasing burden o n diagnostic facilities, so tests simpler than Ziehl-Neelsen staining are n eeded. Such tests should be objective, reproducible, and have at least as g ood a detection limit as 10(4) bacteria/ml. A capture enzyme-linked immunos orbent assay (ELISA) was developed for detection of lipoarabinomannan (LAM) in human sputum samples. As a capture antibody, we used a murine monoclona l antibody against LAM, with rabbit antiserum against Mycobacterium tubercu losis as a source of detector antibodies. The sensitivity of the capture EL ISA was evaluated by using purified LAM and M. tuberculosis whole cells. We were able to detect I ng of purified LAM/ml and 10(4) M. tuberculosis whol e cells/ml, LAM could also be detected in culture filtrate of a 3-week-old culture of M. tuberculosis. The culture filtrate contained approximately 10 0 mu g of LAM/ml, The detection limit in sputum pretreated with N-acetyl-L- cysteine and proteinase K was 10(4) M, tuberculosis whole cells per ml. Thi rty-one (91%) of 34 sputum samples from 18 Vietnamese patients with tubercu losis (32 smear positive and 2 smear negative) were positive in the LAM det ection assay. In contrast, none of the 25 sputum samples from 21 nontubercu lous patients was positive, This specific and sensitive assay for the detec tion of LAM in sputum is potentially useful for the diagnosis of tuberculos is.