Detonation Nanodiamonds for Rapid Detection of Clinical Isolates of Mycobacterium tuberculosis Complex in Broth Culture Media

• Published in: Analytical chemistry (Washington) 2012-09, Vol.84 (18), p.7972-7978
• Main Authors: Soo, Po-Chi, Kung, Ching-Jen, Horng, Yu-Tze, Chang, Kai-Chih, Lee, Jen-Jyh, Peng, Wen-Ping
• Format: Article
• Language: English
• Subjects: Analytical chemistry; Bacterial Proteins - analysis; Biomarkers - metabolism; Cell culture; Chemistry; Culture Media - metabolism; Diamonds; Dispersion; Exact sciences and technology; Gram-positive bacteria; Humans; Mass spectrometry; Miscellaneous; Mycobacterium tuberculosis - isolation & purification; Mycobacterium tuberculosis - metabolism; Nanodiamonds - chemistry; Spectrometric and optical methods; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tuberculosis - microbiology
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac301767z

Routinely used molecular diagnostic methods for mycobacterium identification are expensive and time-consuming. To tackle this problem, we develop a method to streamline identification of Mycobacterium tuberculosis complex (MTBC) in broth culture media by using detonation nanodiamonds (DNDs) as a platform to effectively capture the antigen secreted by MTBC which is cultured in BACTEC MGIT 960, followed by the analysis of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). The 5 nm DNDs can capture the MTBC secretory antigen without albumin interference. With on diamond digestion, we confirm the DND captured antigen is cell filtrate protein 10 (CFP-10) because its Mascot analysis shows a score of 68. The dot blotting method further verifies a positive reaction with anti-CFP-10, indicating that CFP-10 is secreted in the medium of mycobacterium growth indicator tube (MGIT) and captured by DNDs. The minimal CFP-10 protein detection limit was 0.09 μg/mL. Furthermore, our approach can avoid the false-positive identification of MTBC by immunological methods due to cross-reactivity. Five hundred consecutive clinical specimens subjected to routine mycobacteria identification in hospital were used in this study, and the sensitivity of our method is 100% and the specificity is 98%. The analysis of each MTBC sample from culture solution can be finished within 1 h and thus shortens the turnaround time of MTBC identification of gold standard culture methods. In sum, DND MALDI-TOF MS for the detection of MTBC is rapid, specific, safe, reliable, and inexpensive.