WHO Recommended SOPs for Xpert MTB/RIF Sample Processing

The following Standard Operating Procedures (SOPs) are recommended when processing the following extrapulmonary samples:

SOPs for Lymph Node and Tissues Samples

1. Use sterile scissors and forceps to cut-up tissue sample and add 2 ml of sterile Phosphate-buffered Saline (PBS).
2. Grind in sterile mortar and pestle.
3. Transfer approximately 0.7 ml of the homogenized tissue sample to a sterile, conical, screw-capped tube using a transfer pipette.
4. Add a double volume of Cartridge-based Nucleic Acid Amplification Test (CBNAAT) sample reagent (1.4 ml) to 0.7 ml of homogenized tissue using a transfer pipette.
5. Vigorously shake 10 to 20 times or vortex for at least 10 seconds.
6. Incubate for 10 minutes at room temperature, and again shake the specimen vigorously 10 to 20 times or vortex for at least 10 seconds.
7. Incubate the sample at room temperature for an additional 5 minutes.
8. Using a fresh transfer pipette, transfer 2 ml of the processed sample to the CBNAAT cartridge.
9. Load the cartridge into the CBNAAT instrument as per the manufacturer’s instructions.

SOPs for Cerebrospinal Fluid (CSF) Samples

Sample preparation for 5 ml CSF sample:

• Transfer CSF into conical tube and centrifuge (3000 g:15 min).
• Decant supernatant into disinfectant solution (Biosafety Cabinet (BSC) required).
• Resuspend sediment to a volume of 2 ml using CBNAAT sample reagent.
• Add concentrated specimen to the cartridge and proceed with CBNAAT testing.

Sample preparation for 1 - 5 ml CSF sample (including blood-stained samples): 

• Add an equal volume of CBNAAT sample reagent.
• Add 2 ml into the cartridge and proceed with CBNAAT testing.

Sample preparation for 0.1 - 1 ml CSF sample:

• Makeup the volume to 2 ml by adding sample reagent and proceed with CBNAAT testing.

Sample preparation for <0.1 ml CSF sample:

• Insufficient sample for CBNAAT testing.

Each laboratory must optimize this SOP to minimize the error rate.

Resources

• Xpert MTB/RIF Assay for the Diagnosis of Pulmonary and Extrapulmonary TB.

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