SOP for Preparation and Sterilization of Microbial Media

OBJECTIVE

To lay down a procedure for the preparation and sterilization of microbiological media using pre-formulated / dehydrated media.

SCOPE

This SOP is applicable to all the media used for Microbiological testing in PharmaInfo Limited.

RESPONSIBILITY

Microbiologist: Responsible for preparation of media.

Section In charge: To review the procedure.

QC-Head: Responsible for implementing the procedure.

QA Head: Overall compliance

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PROCEDURE

Receipt of Microbial media:

• All microbial media shall be procured from any one of the reputed media manufacturers mainly from the following sources.

1. Hi-Media laboratory
2. Difco Labs
3. Merck

• All media shall be received in the form of dehydrated media or ready-prepared media.
• On receipt of the microbial media ensure that the Certificate of Analysis is received along with each media and verify the media details like lot number, Manufacturing date, and Expiry date in COA with that the label information on the media container.
• If media details match then receive the media, if media details are not match reject the media containers and return to the supplier.
• Enter the received media in Annexure I (Media inward and stock record).
• Allot the In-house ID number for all received dehydrated media containers.
• Media in-house ID shall contain the details like “X-DM/YYY/ZZ”.

   Where 

      X stands for Media code

      DM stands for Dehydrated Media

      YYY stands for serial number shall start from 001

      ZZ stands for the last two digits of the current year.

For example, the media in-house ID for Soya bean Casein Digest Agar will be SCDA-DM/001/21

Where SCDA is the media code & DM is Dehydrated Media, 001 is the serial number and 21 is the last two digits of the current year.

• Refer to Table I in Annexure IV for Media Codes of all media.
• Label each container with the label mentioned in Annexure III.
• Store all the received media as per manufacturer instructions.
• Before using the media for regular analysis perform the Growth Promotion Test as per the current version of SOP for each lot.

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Preparation of Media:

• Follow the manufacturer's instructions for the preparation of all media.
• Whenever the media is taken for a growth promotion test or for regular analysis enter the details of the container ID number in Annexure I.
• Weigh the required quantity of the dehydrated media into a cleaned and dried suitable container which is labeled with the media name and date of preparation.

• Add an adequate quantity of freshly collected purified water.
• Stir/mix properly to get uniform dispersion of media with water.
• Boil the prepared media on a Hot plate to dissolve the media completely.
• The pH can be checked as per the label claim of the Manufacturer.
• Check the pH using a pH meter before and after sterilization. Adjust the pH, as required, by using 1M/0.1 M HCl or 1M/0.1M NaOH before sterilization as that after sterilization pH shall come within the required pH, if pH after sterilization is not within the limit discard that media and prepare freshly for analysis.
• Add supplements for specific media as per Table I in Annexure IV.
• Media shall be prepared on a daily basis based on the samples to be analyzed and prepared media shall be used on the same day.
• Allot the in-house ID number for all prepared media.
• Media in-house ID shall contain the details like “X-PM/YYY/ZZ”.

Where 

     X stands for Media code 

     PM stands for Prepared Media

     Y stands for serial number shall start from 001 

     ZZ stands for the last two digits of the current year.

For example, the media In-house ID for Soya bean Casein Digest Agar will be SCDA-PM/001/21

Where… SCDA is the media code, PM is prepared medium, 001 is serial number and 21 is the last two digits of the current year.

If the same day SCDA is prepared in the second autoclave load the number will be SCDA- PM/002/21.

• Refer to Table I in Annexure IV for Media Codes of all media.
• Dispense the prepared media into individual containers [flasks, tubes] as required.
• Close the containers using cotton plugs and cover the cotton plug with aluminium foil.
• Label the media flasks with the media name and date of preparation.
• Sterilize the media in a steam sterilizer Horizontal Autoclave at 121°C for 15 min, or as directed in Annexure IV Table II.
• Dispense the sterile media as required into sterile containers (Petri plates and tube) at approximately 40°C and allow it to solidify.
• Label each petri plate and test tube with the In-house ID of the medium & date of preparation and use them for regular analysis.
• For prepared solid media (each autoclaved lot), a growth promotion test shall be performed by means of streaking with all organisms mentioned in the current version of SOP by rotating weekly with one organism.

• For prepared liquid media (each autoclaved lot), a growth promotion test shall be performed by means of observing turbidity with all organisms mentioned in the current version of SOP by rotating weekly with one organism.
• The leftover media after completion of analysis shall be disposed of as per the current version of SOP.

ALSO READ: SOP for Growth Promotion Test

Preparation of 1M/ 0.1M HCl and 1M/0.1M NaOH

1M HCl: Dilute 85 mL of conc. HCl was diluted to 1000 mL with purified water to obtain 1M HCl.

0.1M HCl: Dilute 10 mL of 1M HCl to 100 mL to obtain 0.1M HCl solution.

1M NaOH: Dissolve 40 g NaOH in purified water and make it 1000 mL to obtain 1M NaOH.

0.1M NaOH: Dilute 10 mL of 1M NaOH solution to 100 mL to obtain 0.1MNaOH solution.

Precautions during media preparation and usage:

• Label the container with the opened-on date when opened for the growth promotion test / for regular usage.
• Close the container tightly immediately after use.
• Clean the spillage, if any, and immediately sanitize the surface with 70% IPA.
• Use hand gloves and nose masks while handling the dehydrated media.
• Use cleaned and dried glassware for media preparation.
• Care should be taken not to overheat the media, and avoid re-melting the prepared media.
• Avoid excessive shaking of the prepared media to minimize froth/air bubbles.
• Before use, check pre-incubated agar plates/tubes for un-even spread/drying or any contamination.
• All prepared media must be sterilized within 4 hrs.
• Any spillage/breakage in the sterilizer must be cleaned immediately.

Media Reconciliation procedure:

• The fill weight of dehydrated media will differ up to ± 2 % from the stated weight on the bottle.
• Daily media consumption shall be entered in Annexure II.
• After completion of each dehydrated media, the container checks for the less or excess quantity by comparing the quantity mentioned in the consumption record.
• Document the excess or lesser quantity that differs from the consumption record in Annexure II.
• If excess quantity is observed that quantity shall be added to the next media bottle and the same shall be documented in the consumption record in Annexure II.
• If less quantity is observed take a sufficient quantity of media from the next bottle to complete the media preparation and the same shall be documented in the consumption record in Annexure II.
• After completion of each dehydrated media container, reconcile in the stock record (Annexure – I).

ABBREVIATIONS

QC: Quality Control

QA: Quality Assurance

HCl: Hydrochloric Acid

IPA: Iso Propyl Alcohol.

NaOH: Sodium Hydroxide

COA: Certificate of Analysis

SOP: Standard Operating Procedure

ANNEXURE

Nil

REVISION HISTORY

Nil

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