Standard Operating Procedure Title: Preparation of Media For sterility Testing.

Purpose:
• It is established for the preparation of media for sterility test first assure that all the glassware properly washed and sterilized in oven at 200o C for 1 hour.Cool the glassware at room temperature by covering the mouth of glassware from environment.
2. Scope:
• It is applicable to Microbiological Lab.
3. Responsibilities:
• Quality Control Manager
• Microbiologist
4. Procedure:
• FOR FTM:
• Take a sterilized 500 ml screw capped conical flask and take 500 ml of distilled water in it
• .warm it about 70-80oC.
• Weigh 14.750 g of media on weighing boat /tulip funnel and pour it into flask containing distilled water.
• Swirl it to dissolve while heating until a clear solution is obtained.
• Take sterilized 250 ml media glass bottles and pour 100 ml of media in each bottle.
• Autoclave the media bottles at 121oC at apressure of 15 pound.
• After autoclaving cool the media bottles to room temperature.
• Preserve the media for 3-4 days and assure that media is free from contamination.
For SDB:
• Take a sterilized 500 ml screw capped conical flask and take 500 ml of distilled water in it
• .warm it about 40-45oC.
• Weigh 15.0 g of media on weighing boat /tulip funnel and pour it into flask containing distilled water.
• Swirl it to dissolve, until a clear solution is obtained.
• Take sterilized 250 ml media glass bottles and pour 100 ml of media in each bottle.
• Autoclave the media bottles at 121oC at and pressure of 15 pound for 30 minutes.
• After autoclaving cool the media bottles to room temperature.
• Preserve the media for 3-4 days and assure that media is free from contamination.
Peptone water buffer:
• Take a sterilized 500 ml screw capped conical flask and take 500 ml of distilled water in it
• Add 0.5 g of peptone water buffer and shake to dissolve.
• Autoclave the buffer bottles at 121oC at and pressure of 15 pound pressure for 30 minutes.
• After autoclaving cool the buffer to room temperature.
• Preserve the media for 3-4 days and assure that media is free from contamination.
Procedure:
In case of liquid, use 20 representative containers.
• Clean the exterior surface of ampoules with Isopropyl alcohol and gain access to contents in aseptic manner.
• Remove the liquid from test container with sterile pipette or sterile syringe and needle.
• Aseptically transfer 20 ml of the contents from 20 containers to a 200 ml sterile peptone buffer solution.
• Sterilize filter paper, filtration assembly, pipes dress, waste bottle.
• Fit the filtration assembly on waste bottle. Dip one end of the pipe from filtration assembly into peptone buffer solution.
• Filtration assembly is attached with the vacuum pump. Apply vacuum to affect filtration.
• After filtration, Open aseptically remove filter paper containing expected counts with the help of a sterile forcep and cut it into two pieces with the help of a sterile scissor.
• Place half paper aseptically in FTM media and second in SDB media. Incubate FTM media at 30-35oC and SBD media at 20-25oC for seven days.
Interpretation of results:
If FTM is hazy or turbid, bacterial count is positive.
If SDB is hazy or turbid, fungal count is positive.
If both are transparent, bacteria and fungus are absent.
Procedure:
In case of tablet take 20 units, corresponding to not less then 300 mg from each container being tested or to the entire contents of each vial contains less than 300 mg of solids. Clean the exterior of vial with Isopropyl alcohol and gain excess to the contents from container in an aseptic manner. Aseptically transfer about 6.0 g of contents to 400 ml sterile peptone buffer solvent. Sterilize filter, filter holder assembly, pipes, dress and bottle for waste.
Place one end of the pipe from filter holder into the peptone buffer solution.2nd end of the filtration assembly is attached with the vacuum pump apply vacuum to affect filtration, after filtration pass 400 ml of buffer, open aseptically and remove filter paper containing expected count with the help of a sterile forceps and cut it into two pieces with the help of a sterile scissors, place half paper aseptically in FTM media and 2nd in SDB media, Incubate FTM media at 30-35oC SDB 20-25oC for fourteen days.
Interpretation of results: If FTM is hazy or turbid bacterial conc. +ve
If SDB is hazy or turbid fungus +ve
If both are transparent bacterial fungus both assent.

Quality Record(s)/Form(s)

The following quality records shall be generated in accordance with the procedure for Control of Company Quality Records (4.12).

Required Record Form No.
Report of Pyrogen Test QF/059/01