The PaxView TB/NTM MPCR-ULFA Kit, which targets the IS6110 and mtp40 genes for Mycobacterium tuberculosis (MTB) detection, is a novel tool that substitutes gel electrophoresis for universal lateral flow assays. The sensitivity and specificity of this method were compared with those of established methodologies using Indonesian clinical isolates. In this study, 148 sputum specimens isolated from suspected tuberculosis (TB) carriers were examined to evaluate the performance of the PaxView TB/NTM MPCR-ULFA Kit compared to that of smear microscopy and the Xpert MTB/RIF assay. Out of 148 cases, the rate of TB-positive samples evaluated by different methods was 18.2% (27/148; 95% CI 11.9–24.4) for smear microscopy, 20.3% (30/148; 95% CI 13.8–26.8) for the Xpert MTB/RIF, and 34.5% (51/148; 95% CI 26.8–42.1) for the PaxView TB/NTM MPCR-ULFA Kit. Twenty sputum specimens from healthy subjects were also tested, all of which rendered negative results via the three diagnostic tools examined herein. Compared to the Xpert MTB/RIF, the PaxView TB/NTM MPCR-ULFA Kit was found to possess a 96.7% sensitivity (29/30; 95% CI 90.3-100).Moreover, the PaxView TB/NTM MPCR-ULFA Kit detected 18.6% (22/118, 95% CI 11.6–25.6) of Xpert MTB/RIF MTB negative samples and 20.7% (25/121, 95% CI 13.5–27.9) of smear microscopy negative samples as MTB positive. The PaxView TB/NTM MPCR-ULFA Kit could be a useful molecular diagnostic tool to identify MTB in clinical samples in resource-limited countries, as this procedure is more cost-effective and sensitive than the Xpert MTB/RIF, and more convenient than conventional PCR gel electrophoresis approaches.
The PaxView TB/NTM MPCR-ULFA Kit, which targets the IS6110 and mtp40 genes for Mycobacterium tuberculosis (MTB) detection, is a novel tool that substitutes gel electrophoresis for universal lateral flow assays. The sensitivity and specificity of this method were compared with those of established methodologies using Indonesian clinical isolates. In this study, 317 sputum specimens isolated from tuberculosis (TB) suspects were examined to evaluate the performance of the PaxView TB/NTM MPCR-ULFA Kit compared to that of smear microscopy and the Xpert MTB/RIF assay. Out of 317 cases, the rate of TB-positive samples evaluated by different methods was 33.4% (106/317; 95% CI 28.2-38.6) for smear microscopy, 37.9% (120/317; 95% CI 32.5-43.2) for the Xpert MTB/RIF, and 40.7% (129/317; 95% CI 35.3-46.1) for the PaxView TB/NTM MPCR-ULFA Kit. Compared to the Xpert MTB/RIF as a standard reference, the PaxView TB/NTM MPCR-ULFA Kit was found to possess a 92.5% sensitivity (111/120; 95% CI 87.8-97.2), a 90.8% specificity (179/197; 95% CI 86.8-94.8), 86.0% PPV (111/129; 95% CI 80.0-92.0), and a NPV 95.2% (179/188; 95% CI 92.2-98.3). The PaxView TB/NTM MPCR-ULFA Kit could be a useful molecular diagnostic tool to identify MTB in clinical samples in resource-limited countries, as this procedure is more cost-effective and sensitive than the Xpert MTB/RIF, and more convenient than conventional PCR gel electrophoresis approaches.
Ziehl-Neelsen Smear Microscopy
Ziehl-Neelsen direct AFB smear and grading was performed by the technicians from each institute. Briefly, smears are fixed on heated surface (60 °C for at least 10 minutes) and then flooded with carbolfuchsin (primary stain). Smears then are heated to almost boiling and are after the smears were allowed to sit for 5 min, then the slides are washed in distilled water.
The slides were then decolorized with 3% HCI in 95% ethanol for approximately 1 minute and washed with water. The methylene blue (counterstain) was then flooded to the slide and allowed to sit for 1 min before the slides being washed with distilled water and let dry upright. The slides were then examined microscopically according to the International Union Against Tuberculosis and Lung Disease (IUATLD) method9.DNA extraction from specimens After SM, the remaining sputum specimen was treated with Xpert MTB/RIF buffer. After proceeding Xpert MTB/RIF test, the remaining buffer treated sputum were stored in the refrigerator until maximum 20 days before DNA extraction from the TB bacilli for PaxView TB/NTM MPCR-ULFA test.
DNA were extracted by the PaxView DNA Extraction Kit (PaxGenBio, Korea). Briefly, 1000 μl of the specimen pretreated with the Xpert MTB/RIF buffer was transferred into 1.5 ml screw capped tubes and then centrifuged at 13,000 rpm for 3 min. After the supernatant was discarded, 1000 μl of washing buffer were added (provided by the PaxView DNA Extraction Kit). This mixture was then centrifuged again at 13,000 rpm for 3 min, after that the supernatant was discarded. After washing the specimen once more, 100 μl of elution buffer were added into the tube, which was then transferred to a 95 °C heating block for 15 min. After centrifugation, 20 μl of supernatant were transferred into a new tube and used as a template.
Description: A polyclonal antibody against NTM. Recognizes NTM from Human. This antibody is Unconjugated. Tested in the following application: WB, IHC, ELISA;WB:1/500-1/2000.IHC:1/100-1/300.ELISA:1/20000
Description: A polyclonal antibody against NTM. Recognizes NTM from Human. This antibody is Unconjugated. Tested in the following application: ELISA, WB;WB:1:500-1:3000
Description: A polyclonal antibody against NTM. Recognizes NTM from Human. This antibody is Unconjugated. Tested in the following application: ELISA, WB;WB:1:500-1:3000
Description: A polyclonal antibody against NTM. Recognizes NTM from Human. This antibody is Unconjugated. Tested in the following application: ELISA, IHC; Recommended dilution: IHC:1:20-1:200
Description: A polyclonal antibody against NTM. Recognizes NTM from Human, Mouse. This antibody is Unconjugated. Tested in the following application: ELISA, WB, IHC; Recommended dilution: WB:1:1000-1:5000, IHC:1:20-1:200