Shreffler:Aseptic Technique

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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