fa Original article Original article <div dir="ltr" style="text-align: justify;"><strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">Introduction:</span></span></span></strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;"> Extracellular vesicles (EVs) are bacterial products with diverse biological roles. Like many microorganisms, <em>Mycobacterium</em> <em>kansasii</em> as a nontuberculous mycobacteria (NTM), can naturally release EVs. The aim of the present study was the extraction and biological evaluation of <em>M. kansasii</em> as a vaccine candidate against <a name="_Hlk494623320">mycobacterial pulmonary infections</a>.</span></span></span> <strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">Methods: </span></span></span></strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">After bacterial culture of the standard species of <em>M. kansasii</em>, the EVs extraction was done by density gradient ultra-centrifugation method and biological evaluations of EVs were performed by SDS-PAGE and electron microscopy. Endotoxin safety of the EVs was evaluated by LAL test. </span></span></span><strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">R</span></span></span><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">esults:</span></span></strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;"> SDS-PAGE result showed more than 5 prominent protein bands (60-180 kDa). The intactness of the vesicles was verified by electron microscopy through which the spherical configuration of EVs with diameter of 200-300 nm could be observed. The amount of lipopolysaccharide (LPS) contamination existing in EVs was in the specified application range of biological products. </span></span></span><strong><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">Conclusion: </span></span></strong><span style="letter-spacing:-.1pt;"><span style="font-family:times new roman,serif;"><span style="font-size:10.0pt;">EVs were prepared with acceptable quality composition with intact conformational structure throughout the extraction procedure. The extracted EVs had the initial requirements as an immunogenic molecule, such as safety, stability, inexpensiveness and antigens possession which based on the similarities between <em>M. kansasii</em> and <em>M. tuberculosis</em>, make them a suitable candidate for future prophylactics, therapeutic, detection and adjuvants studies against <a name="_Hlk494623368">mycobacterial pulmonary infections</a></span></span></span><span style="font-family:times new roman,serif;"><span style="font-size:11.0pt;">.</span></span></div> Extracellular vesicles (EVs), Mycobacterium kansasii, adjuvant. 19 22 http://vacres.pasteur.ac.ir/browse.php?a_code=A-10-1-36&slc_lang=fa&sid=1 Z Hoseini Tavassol No Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, I. R. Iran. F Vaziri No Department of Mycobacteriology and Pulmonary Research, Pasteur Institute of Iran, Tehran, I. R. Iran./ Microbiology Research Center, Pasteur Institute of Iran, Tehran, I. R. Iran. SD Siadat Yes Department of Mycobacteriology and Pulmonary Research, Pasteur Institute of Iran, Tehran, I. R. Iran./ Microbiology Research Center, Pasteur Institute of Iran, Tehran, I. R. Iran.