Syringe release of Toxoplasma

  1. Start with a highly infected monolayer (e.g. 0.5 - 1 mL of RH on a T25 overnight)
  2. (Larger than T25 only): Aspirate majority of media (all but 3 - 10 mL, depending on size of flask)
  3. Scrape cells from flask with a sterile rubber scraper, e.g. autoclaved Fisher #22261768 (or disposable if you are doing immune cell culture)
  4. Attach sterile 27 gauge blunt needle (SA Infusion technologies #B27-100) to syringe
    • For T25: Use 3 mL syringe directly inserted into flask and pass contents of flask through syringe 2-3x.
    • For T75 or larger: Pipette media and cell remnants (and parasites!) into open syringe, use plunger to force cell debris through needle and into conical tube.
  5. Dispose of empty syringe/needle in sharps container
  6. Sediment parasites and cell debris in TC centrifuge (~100 - 150 RCF)
  7. Aspirate media
  8. Optionally wash with media and repeat spin.
  9. Bring up parasites in appropriate media or buffer and continue with experiment (e.g. pass parasites).

Note: This is often (incorrectly) referred to as “syringe lysis” of parasites. Since it is the host cells that are being disrupted, that is a misnomer.

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