This post was most recently updated on April 23rd, 2015
Tran-isolate medium is used to inoculate and transport CSF sample, If the CSF cannot be transported to a microbiology laboratory immediately (within 1 hour from the time of collection) for culture and analysis of cerebrospinal fluids from patients with bacterial meningitis.
Trans-Isolate medium is a diphasic medium, consisting:
- Slant: Charcoal-starch agar
- Broth: Soybean-casein digest-gelatin broth buffered at pH 7.2 with 0.1 M 3-(N-morpholino) propanesulfonic acid buffer.
T-I medium supports the growth and survival of Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenza. In case of stock culture T-I medium supported the growth for at least 3 months.
Procedure for primary culture from Trans-Isolate (T-I) medium
- Remove the aluminum cap of Trans-isolate medium with forceps
- Wipe the rubber stopper with a 70% alcohol swab (do not use povidone-iodine as it may be carried into the medium by the passing needle, thus inhibiting the growth of bacteria.)
- Aseptically inoculate 0.5-1.0 ml of the CSF into the T-I medium with the use of syringe
- Incubate the inoculated T-I medium at 35-37°C with ~5% CO2 (or in a candle-jar) overnight or until transport is possible
- If the T-I medium can be transported to a microbiology laboratory the same day of inoculation, do not vent the T-I bottle until it arrives in the receiving laboratory. Upon arrival, wipe the rubber stopper with 70% alcohol, insert a venting needle into the T-I bottle, incubate at 35-37°C with ~5% CO2 (or in a candle-jar), and observe daily for turbidity in the liquid phase for up to 7 days.
- Prior to subculture, remove the venting needle and wipe the rubber stopper with 70% alcohol.
- Use a sterile needle and syringe to transfer 50-100 µl of the liquid portion of the T-I medium onto both a Blood agar Plate (BAP) and Chocolate agar (CAP) for primary culture.
- Approximately 50-100 µl is used to streak each plate. To streak two plates, draw approximately 100-200 µl with the syringe at one time to minimize the possibility of contaminating the T-I medium.
- Streak the BAP and CAP for isolation, incubate the plates at 35-37°C with ~5% CO2 (or in a candle-jar), and examine the plates daily for up to 72 hours.
- If no growth is observed, subculture the T-I medium again on day 4 and day 7.
- Isolates should always be inspected for purity of growth by looking at colony morphology before any testing is performed. If any contamination is seen, cultures should be re-streaked to ensure purity prior to testing.