Hello Hafid,

We use NaOH for 15 minutes followed by an indicator buffer to bring the specimen back to pH 7 once decontaminated. We find that if the tissue is cut up finely enough with a scalpel (under strict safety protocols), the specimen will be sufficiently decontaminated. However, you will always have a number of specimens that are grossly contaminated with bacteria/fungus that will need re-decontamination at some point. The BD MGIT medium enriches for all organisms unfortunately (I assume you are using the MGIT 960 system). We re-decontaminate for 30 minutes if required. The timing of decontamination is a fine balance between killing all organisms (including the mycobacteria) and just killing the bacteria/fungus (and keeping the mycobacteria alive).

Hope this is helpful,

Emma Roycroft.

Thank you Emma,  It's exactly what we are doing NaOH for 15 min, and redecontaminate for further 15 min, but we have still some contaminants.

We will try to incubate for more time, even if we loose some bacilli . Thank you

You could add antibiotic mixture which can kill bacteria / fungus but not inhibit M.tb growth, the final concentration we used is as the following: 40 U/ml polymyxin B, 4 mg/ml amphotericin, 50 mg/ml carbenicillin, 2 mg/ml trimethoprim.

Hope this is helpful to your work.

Thank you for your inputs.

We used PANTA before, but it's time to order it. We are processing those tissus for research project to describe M. xenopi infection in birds (that is rarely reported). I am aware of treating with NaOH for more time to overkill mycobacteria since M. xenopi grows slowly. Thank you