1.0           OBJECTIVE:

To lay down a standard procedure for preparation and sterilization of microbiological culture media.

2.0           SCOPE:

This Standard Operating Procedure is applicable for media preparation and sterilization in Microbiology laboratory of Quality Control Department.

3.0           RESPONSIBILITY:

                Microbiologist shall be responsible to follow the procedure mention in this SOP.


Department Head & QA Head shall be accountable for compliance of this SOP.

5.0           ATTACHMENTS:

Media preparation record                                                                  –Attachment-I

6.0           PROCEDURE:

6.1           General precautions:

6.1.1        Take clean, dried glass conical flask/borosilicate glass bottle of desired volume as per quantity of


6.1.2        Use a separate spatula for separate media to avoid cross contamination.

6.1.3        Clean the balance in between successive weighing.

6.2           Preparation and sterilization:

6.2.1        Use dehydrated culture media from Merk, Hi Media Laboratories Ltd or any of the Authentic Source.

Check the in house use before date prior to use of the media.

6.2.2        Take clean dried conical flask/borosilicate glass bottle of desired volume as per quantity of media.

Transfer the half of the volume of purified water of required quantity in the conical flask.

6.2.3        Weigh required quantity of de-hydrated media as per volume required, transfer in conical flask and

re-hydrate it as per manufacturer instructions. Make up the volume with remaining quantity of water.

6.2.4        Mix well and heat gently to dissolve the media. Note down the weight taken for each medium.

6.2.5        Check the pH of the media before sterilization. Adjust the required pH (specified on the respective

medium pack) by using either 0.1 M HCl or 0.1 M NaOH solution if required.

6.2.6         In case of broth medium, distribute the suitable quantity of the broth medium in appropriate glass

bottle or test tubes.

6.2.7         Plug the bottles with screw cap or test tubes with a cotton plug or screw cap, wrap with aluminum


6.2.8         Sterilize the media by autoclaving at temperature121°C and 15 psi/kg/cm² pressure for15 minutes or

as per manufacturer instructions.

6.2.9          After sterilization, use the media for desired purpose and Store the sterilized broth media below 25°C

till further use.

6.2.10       For agar media, after auto claving, cool the medium to about 45-50°C then pour the media

aseptically into sterile Petri plates in 15 to 20 ml quantity in each plate. Allow the plates to solidify.

6.2.11       Close the plates invert them and incubate them for desired purpose.

6.2.12       After preparation of media, record the following details in the format of media preparation record in

format no. SOP/ML/005/F01-00.

6.2.13       Name of media, Batch No., Use before, Date of preparation, Total weight taken, Total volume, pH

before sterilization and  Prepared by.

6.2.14       All the media prepared in a day should be assigned a respective batch no. and these batch nos.

should be entered in the media preparation, utility & destruction record.

6.3            Assigning batch/lot number:

6.3.1          All the media prepared in one autoclaved cycle should have respective batch numbers and these

batch numbers should be entered in media preparation record.

6.3.2         Batch no. of autoclaved media should be allotted in the following manner:


Where XXX : Autoclave load no. starting from 001 for every year

And ZZZZZZZ: Batch no. of media provided by the manufacturer.

6.4            Storage of rehydrated sterilized media:

6.4.1         Broth medium      : Stored below 25°C

6.4.2         Molten agar media    : Stored between 55-60°C

6.4.3         Prepared agar plates : Stored below 25°C

7.0           REFERENCES:



SOP : Standard operating procedure

°C   : Degree Celsius

Psi   : Pound per square inch

Ml    :  Milliliter

HCl :  Hydrochloric acid

NaOH         : Sodium Hydroxide

CC No.        : Change Control number

QA              : Quality Assurance

QC  : Quality Control


Quality Assurance Department

Quality Control Department


Version Number Effective Date Reason for Revision
00 20/04/17 New SOP


Title : Media preparation record
Version No. : 00 Page No. :  1 of 1


Effective Date :   Review Date :  


S. No. Date Name of Media Media Quantity taken Volume prepared (ml) pH before sterilization Autoclave lot. no./Media batch/lot no. Prepared by Checked by




Microbiology Labs Documents