SOP For Media Preparation used for microbiological testings is described in this post which you can follow in the section of microbiological lab.
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Objective:
This procedure has been established for preparing culture media i.e. [FTM(Fluid Thioglycolate Media), TSB (Tryptic Soy Broth), PW(Peptone Water) and Nutrient Agar] used for microbiological testing's.
Scope:
This procedure is applicable to microbiology lab.
Responsibility:
Microbiologist.
Microbiology Lab Assistant.
Accountability:
Quality Control Manager.
Procedure:
FTM (Fluid Thioglycolate Media):
For preparing 100ml of media (if the manufacturer is MERCK*)weigh accurately 3g of the powder media in a calibrated weighing balance in a clean and dry beaker and transfer the powder in the media bottle and makeup the final volume upto 100ml in a calibrated measuring cylinder.
Switch "ON" the hot plate and set the temperature at 80oC and place the media on heating and place the magnetic stirrer in the media bottle for continuous mixing and switch "ON" the magnetic stirrer.
before heating the color of the solution will be brown which on heating must be converted to pink color when it changes its color from brown to pink remove the media bottle from heating and remove the magnet from the solution and place the cap over the bottle and move the media for steam heat sterilization (autoclavation) at 121oC/15psi/15minutes. After sterilization cool the media and place in hot incubator at a temperature of 32oC±2.5oC atleast for 48hrs before using for sterility test.
TSB (Tryptic Soy Broth):
For preparing 100ml of media (if the manufacturer is MERCK*)weigh accurately 3g of the powder media in a calibrated weighing balance in a clean and dry beaker and transfer the powder in the media bottle and makeup the final volume upto 100ml in a calibrated measuring cylinder.
Switch "ON" the magnetic stirrer and place the media for continuous mixing. At the end of homogeneous solution remove the magnet from the solution and place the cap over the bottle and move the media for steam heat sterilization (autoclavation) at 121oC/15psi/15minutes.
After sterilization cool the media and place in cool/mold incubator at a temperature of 22oC±2.5oC atleast for 48hrs before using for sterility test.
PW(Buffered Peptone Water):
For preparing 1liter of media (if the manufacturer is MERCK*)weigh accurately 25.5g of the powder media in a calibrated weighing balance in a clean and dry beaker and transfer the powder in the media bottle and makeup the final volume upto 1liter in a calibrated measuring cylinder.
Switch "ON" the magnetic stirrer and place the media for continuous mixing. At the end of homogeneous solution remove the magnet from the solution and place the cap over the bottle and move the media for steam heat sterilization (autoclavation) at 121oC/15psi/15minutes.
After sterilization cool the media and place in cool/mold incubator at a temperature of 22oC±2.5oC atleast for 48hrs before using for sterility test.
Nutrient Agar:
For preparing 1liter of media (if the manufacturer is MERCK*)weigh accurately 28g of the powder media in a calibrated weighing balance in a clean and dry beaker and transfer the powder in the media bottle and makeup the final volume upto 1liter in a calibrated measuring cylinder.
Switch "ON" the hot plate and set the temperature at 120oC and place the media on heating and place the magnetic stirrer in the media bottle for continuous mixing and switch "ON" the magnetic stirrer.
At the end of clear/transparent homogeneous solution remove the magnet from the solution and place the cap over the bottle and move the media for steam heat sterilization (autoclavation) at 121oC/15psi/15minutes.
After sterilization cool the media and place in hot incubator at a temperature of 32oC±2.5oC atleast for 48hrs before using for sterility test.
Plate Pouring:
Sterilize (Dry Heat Sterilization) at a temperature of 180oC for 2.5hrs and then move the sterilize nutrient agar media and glass petri-plates to sterile area microbiology lab through pass box under laminar flow cabinet.
Aseptically pour 20ml-25ml of the sterile, molten agar medium, cooled to 40 to 45°C, in each Petri dish.
Allow the medium to solidify and pre-incubate at the desired temperature for 48hrs before use.
Prepared Petri dishes, if not used immediately shall be used within 15 days of preparation.
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