SOP for serial dilution of microbial culture is described in this post which you can follow in the section of microbiological lab.


Objective:

This procedure has been established for preparing or diluting the microbial culture by serial dilution method.

Scope:

This procedure is applicable to microbiology lab.

Responsibility:

Microbiologist.                             

Accountability:

Quality Control Manager.            

Procedure:   

For refreshing the microbial culture on weekly basis transfer a loopful of microbial culture to 10ml tryptic soy broth (TSB) and pour 20-25ml of the nutrient agar and allow solidifying.

Incubate the bacterial culture at 32.5 ± 2.5°C for 2-3days and fungal culture at 22.5 ± 2.5°C for 3-5days

Then at the end of incubation period serially dilute the (Optimum growth of culture obtained) in 0.9% normal saline solution previously sterilized in a ratio of 1: 10 dilution i.e. (1ml of the culture diluted in 10ml of the solvent) so that the dilution must yield atleast a count of 10–100 CFU/ml.

Verification of CFU/ml:

Count the number of CFU/ml from freshly prepared dilution of the culture by transferring 1ml of the inoculum on nutrient agar plate for Bacteria and on Sabouraud dextrose agar plate for Fungi by pour plate method and incubate the plates at their respective time and temperatures.

After the completion of incubation period count the number of colonies observed on plate y colon counter & record the average number of CFU in respective record form.

At the end calculate CFU/ml by comparing the data i.e. (ml of dilution used for test and number of observed CFU by that dilution).

Storage & Usage:

Store the freshly prepared microbial culture at 2°C-8°C in refrigerator for one

week and use this culture within one week after one week dispose of this 

culture as per SOP of disposal and record in logbook.