Detonation nanodiamonds for rapid detection of clinical isolates of Mycobacterium tuberculosis complex in broth culture media.
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.
Department of Physics, National Dong Hwa University, Shoufeng, Hualien, Taiwan, ROC.
Po-Chi
PC
Soo
Ching-Jen
CJ
Kung
Yu-Tze
YT
Horng
Kai-Chih
KC
Chang
Jen-Jyh
JJ
Lee
Wen-Ping
WP
Peng
D001426:Bacterial Proteins / Q000032:analysis; D015415:Biomarkers / Q000378:metabolism; D003470:Culture Media / Q000378:metabolism; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000302:isolation & purification* / Q000378:metabolism; D058612:Nanodiamonds / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014376:Tuberculosis / Q000382:microbiology
D001426:Bacterial Proteins; D015415:Biomarkers; D003470:Culture Media; D058612:Nanodiamonds