Ohio Water Microbiology Lab
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OWML: QA/QC
Quality Assurance/Quality Control Manual: Ohio Water Microbiology
Laboratory
Autoclaves
Sterilization
is the process that eliminates living organisms from substances or objects. The
Ohio Water Microbiology Laboratory (OWML)
is equipped with three autoclaves for sterilization of glassware, reagents,
media, and disposables—two medium-sized autoclaves (Market Forge) that are
operated in the side laboratory and one large autoclave (Consolidated) that is
operated in the warehouse.
- Dishes that need to be
sterilized are wrapped in aluminum foil or kraft paper and placed in the
autoclave for moist heat sterilization. Clean and sterile dishes are stored
in closed cupboards until use.
- The autoclaves are operated at
15 lb/in2 steam pressure, producing an inside temperature of 121 to 124oC
(American Public Health Association, 2005, Section 9020B). Do not overload
the autoclave. Autoclave time depends on the type and amount of equipment as
follows:
- Glassware and up to 250 mL
of liquid—15 minutes
- 500 to 2,000 mL liquid—30
minutes
- Greater than 2,000 mL to
6,000 mL liquid—15 minutes per 1,000 mL
- Greater than 6,000 mL
liquid—90 minutes
- Carbohydrate-containing
media—15 minutes (no more than 250 mL volumes)
- Pathogenic organisms—30
minutes, allow autoclave chamber pressure to decrease, then run for a 60
minute cycle
- Contaminated materials and
discarded cultures—45 minutes
- Operating temperature and
pressure are checked once a week. Heat-sterilizing tape is used with
each run to identify supplies that have been properly sterilized and
checks the performance of the autoclave. The performance is also checked
monthly by using spore indicators and recorded in the LIMS.
- If the autoclave does not
reach the specified temperature or fails the spore indicator test,
service the autoclave and re-sterilize all glassware and reagents that
were insufficiently sterilized.
- For the two medium-sized
autoclaves, general maintenance is as follows:
- The autoclaves are operated
using deionized water.
- At the end of the week,
autoclaves are drained. Twice a month, autoclaves are cleaned with
Liqui-nox, rinsed with water, and drained. The condensate holding tank
is drained daily or as needed. The cleaning date is recorded in the LIMS.
- Twice a year, have a
contractor inspect and calibrate the autoclaves and perform preventive
maintenance. Preventive maintenance dates are recorded in the LIMS.
- Twice a year, clean the
chambers with 10% muriatic acid and flush well with water. Cleaning
dates are recorded in the LIMS.
- For the large autoclave,
general maintenance is as follows:
- Once a month, clean chamber
with water and liquinox. Cleaning dates are recorded in the LIMS.
- Twice a year, have a
contractor perform preventive maintenance and inspection, clean and
service the generator, clean the door gasket and head ring, apply
graphite to the door gasket, oil the door hinge pins, and lubricate the
door hub. Preventive maintenance dates are recorded in the LIMS.
- Twice a year, clean the
chamber with 10% muriatic acid and flush well with water. Cleaning dates
are recorded in the LIMS.
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