When it comes to reading and utilizing CRM certificates, understanding key points is essential for effective usage. The supplier of the certified reference material typically markets these as daily controls, and the certificates are labelled accordingly.

Key points within the certificates

• Storage: Upon receipt, it is crucial to store the products in a dark and cool place (3 - 5 °C).
  Maintaining this condition is necessary for the material to retain the level of target organisms until the expiry date.
• Dilution instructions: The certificate includes information for performing primary dilution of the samples, detailed in Instructions for Use.
• Expiry dates: there are two expiry dates specified:
  • Product Expiry:
    This marks the date when the reference material (freeze dried or powder) expires.
  • Certificate Expiry:
    This is the date when the certificate with specified levels of target organisms expires.
  • Recertification:
    The certificate indicates whether the material will be recertified (resulting in a new certificate), or if it is final and expires simultaneously with the products shelf life. New certificates are issued at regular intervals until the product’s shelf life expires. Upon certificate expiry, a new one will be provided if needed.
  • Assigned value:
    This represents the expected level of the target organism, along with low and high limits, indicating 3 standard deviations.
  • Designation:
    Note the designation on the certificate. Freeze-dried microorganisms in glass vials specify the level per vial (CFU/vial) while the powder samples indicate levels as CFU/g.

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Example of a Reference Material Certificate

1. Material identification: Indicates which material the certificate applies to

2. Batch number: Verify that the batch number matches that of the certificate

3. Microorganisms content: Details the content of the different microorganisms, with traceability to the strains

4. Usage instructions: Instructions for use of the material

5. Expiry dates: Specifies expiry dates for both the certificate and the product, along with information on recertification

6. Measurement unit: Indicates whether the measurement is per vial or gram

7. Assigned value: Lists the expected level of various target organisms

8. High and low limits of the assigned value

Sample preparation

Example of simulated water sample preparation

The vial contains, for example, 4000 CFU/vial of P. aeruginosa, this represents approximately 400 CFU/ml when when diluted 1/10.

If 1 ml of this solution is added to 1 liter of water, the expected level will be around 40 CFU when filtering a 100 ml water sample. If a 10 ml water sample is filtered, approximately 4 CFU will be obtained.

Choose appropriate volume from the vial to achieve the desired concentration.

Example of simulated food sample preparation

The vial contains, for example, 2900 CFU/vial of C. perfringens, this represents approximately 290 CFU/ml when diluted 1/10.

A direct plating from this primary dilution can be perfomed. Pour plating 1 ml of the resuscitated material into TSC agar will result in an overgrown dish of 290 CFU. An expected level of approximately 29 CFU will be obtained by plating 0,1 ml of the material. Alternatively, the same expected level of 29 CFU can be obtained from 1 ml of a -2 dilution.

Choose appropriate volume from the vial to achieve the desired concentration.

Example simulated food sample preparation with matrix

The vial contains, for example, 25 000 CFU/vial of E. coli, this represents approximately 2 500 CFU/ml when diluted 1/10. 

Weigh 10 grams of the selected food matrix sample. Combine 90 ml of dilution fluid and 1 ml of resuscitated material and homogenize. Pour plate 1 ml from this primary dilution into TSC agar, the expected level will be approximately 29 CFU/g.

Choose appropriate volume from the vial to achieve the desired concentration.

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