Shreffler:Aseptic Technique

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General Guidelines

  • Protect eyes, mucous membranes, open cuts and wounds from contact with biohazardous material.
  • Use personal protective equipment as necessary (i.e. gloves, goggles, mask, other protective clothing).
  • Carry out all culture operations under a laminar flow hood.
  • Disinfect all surfaces with a disinfectant solution prior to use.
    • Swab down the working surface liberally with 70% ethanol solution.
    • Periodically spread 70% ethanol solution over the exterior of gloves to minimize contamination.
    • Swab all glassware (medium bottles, beakers, etc.) with 70% ethanol solution before placing them inside the hood.
    • In case of any spill, spread 70% ethanol solution and swab immediately with non-linting wipes.
  • Replace gloves if torn, and discard after use. Do not wear them when entering any other lab area.
  • Bring into the work area only those items needed for a particular procedure.
    • Leave a wide clear space in the center of the hood (not just the front edge) to work on.
    • To prevent blockage of proper air flow and to minimize turbulence, do not clutter the work area.
    • Make sure to have easy access to all items. Do not reach over an item to retrieve another (especially over an open bottle or flask).
  • Never perform mouth pipetting; always use a pipetter.
    • Use sterile wrapped pipettes and discard them into a biohazard waste container after use. Check that the pipette wrapping is not broken or damaged, to confirm sterility.
    • Make sure not to touch the tip of the pipette to the rim of any flask or sterile bottle.
  • Inspect the vessels to be used:
    • T-flask - Must be free of visible contamination or breakage, or lack container identification. The plastic covering the flask must be intact.
    • Bottles - Check for cracks and expiration dates.
    • Spinner flasks - Check for cracks, expiration dates, and proper assembly.
  • When handling sterile containers with caps or lids, place the cap on its side if it must be laid on the work surface.
  • Discard any biohazardous or contaminated material immediately.
  • When finished, clean the work area by wiping with 70% ethanol solution.

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