Step 1: Prepare the Test Product
The customer product is prepared and diluted using a suitable sterile diluent according to the characteristics of the product and applicable USP <61> requirements.

Step 2: Perform Microbial Enumeration
The prepared product is processed using a suitable microbial enumeration method according to the product characteristics and applicable USP <61> requirements. Enumeration methods may include:

• Membrane filtration: The prepared product is passed through a membrane filter to retain microorganisms. The membrane is then transferred to a suitable culture medium for incubation and enumeration.
• Plate-count methods: The prepared product is tested using suitable plate-count techniques, such as the pour plate or spread plate method. Following incubation, visible colonies are counted.
• Most Probable Number (MPN): The prepared product is inoculated into suitable culture media using a series of dilutions. Following incubation, the presence or absence of microbial growth is evaluated, and the microbial count is estimated using the MPN method.

Method selection: The appropriate enumeration method is selected based on the characteristics of the product and its suitability for microbial recovery and enumeration.

Step 3: Incubate the Test Media
The samples are incubated under appropriate temperature and time conditions to allow the recovered microorganisms to grow and form visible colonies.

• Bacteria: Incubated at 30–35°C for 3 days using Tryptic Soy Agar (TSA).
• Yeasts and moulds: Incubated at 20–25°C for 5 days using Sabouraud Dextrose Agar (SDA).

Step 4: Count and Report the Results
The results are reported as colony-forming units (CFU) or the applicable quantitative result obtained from the selected enumeration method.

Results are reported as:

• Total Aerobic Microbial Count (TAMC)
• Total Yeasts and Moulds Count (TYMC)

Acceptance Criteria

Acceptance criteria are evaluated according to USP <1111> and other applicable regulatory requirements. The applicable microbial limits depend on factors such as the product category, route of administration, and intended use.