3M Petrifilm™ Coliform Count Plates Operating instructions

3M Petrifilm™ Coliform Count Plates Operating instructions

Interpretation Guide

The 3M

Petrifilm

Coliform Count Plate is a sample-ready-culture medium system which contains modified Violet Red Bile nutrients, a cold-water-soluble gelling agent and a tetrazolium indicator that facilitates colony enumeration.

CC

Coliform Count Plate

Coliform Definitions by Method

The United States Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) defines coliforms as Gram negative rods, which produce acid and gas from lactose fermentation. Coliform colonies growing on the 3M ™ Petrifilm ™ Coliform Count Plate produce acid, which causes the pH indicator to deepen the gel color, and gas trapped around red colonies. In this interpretation guide, the number of coliforms per the FDA BAM definition is the number of red colonies with gas.

ISO defines coliforms by their ability to grow in method-specific, selective media. ISO method 4832 enumerates typical coliform colonies on Violet Red Bile Lactose (VRBL) agar, with confirmation of atypical colonies. On the 3M Petrifilm Coliform Count Plate, these coliforms are indicated by red colonies with or without gas production. ISO method 4831, enumerating coliforms by the most probable number (MPN) method, defines coliforms by their ability to grow and produce gas in the conditions described in the standard. On the 3M Petrifilm Coliform Count Plate, these coliforms are indicated by red colonies with gas.

It is also possible to enumerate thermotolerant coliforms on the 3M Petrifilm Coliform Count Plate. Typically thermotolerant coliforms can be selected with an elevated incubation temperature. One example of a method for enumeration of thermotolerant coliforms is described in NF V08 060. Reading the total of red colonies on a 3M Petrifilm Coliform Count Plate incubated at 44°C ± 1°C for 24h ± 2h yields results equivalent to enumeration with NF V08 060. Please refer to the product instructions for additional information.

Figure 1 Total colonies with gas = 69 Total colonies = 94

The definition of coliforms may vary by country. Please refer to section above and product instructions for definitions.

Coliform Count Plate Figure 2 No growth = 0

Notice the changes in gel color in Figures 2–5. As the coliform count increases, the gel color deepens. Background bubbles are a characteristic of the gel and are not a result of coliform growth.

2

1

Figure 3 Total colonies with gas = 79 Total colonies = 109

The recommended counting limit on 3M Petrifilm Coliform Count Plates is less than 150. Do not count colonies that appear on the foam barrier because they are removed from the selective influence of the medium (see Circle 1).

Figure 4 Estimated total coliform count = 220

The circular growth area is approximately 20cm 2 . Estimates can be made on plates containing greater than 150 colonies by counting the number of colonies in one or more representative squares and determining the average number per square. Multiply the average number by 20 to determine the estimated count per plate.

For a more accurate count, further dilution of sample may be necessary.

Figure 5 Total coliform count = TNTC

3M Petrifilm Coliform Count Plates with colonies that are too numerous to count (TNTC) have one or more of the following characteristics: many small colonies, many gas bubbles, and a deepening of the gel color.

For a more accurate count, further dilution of sample may be necessary.

Figure 6 Total count = TNTC

When high numbers of non-coliform organisms such as

Pseudomona

s are present on 3M Petrifilm Coliform Count Plates, the gel may turn yellow.

For a more accurate count, further dilution of sample may be necessary.

3

3 2 1

Figure 7 Total colonies with gas = 2 Total colonies = 2

Food particles are irregularly shaped and are not associated with gas bubbles.

Figure 8 Total colonies with gas = 8 Total colonies = 15

Bubble patterns may vary. Gas may disrupt the colony so that the colony “outlines” the bubble (see Circles 1 and 2). Artifact bubbles may result from improper inoculation or from trapped air within the sample. They are irregularly shaped and are not associated with a colony (see Circle 3).

4

Reminders For Use Storage Inoculation

Petrifilm

Coliform

≤ 8°C 1

Store

unopened pouches of plates at <8°C (<46°F). Use before expiration date on package. In areas of high humidity where condensate may be an issue, it is best to allow pouches to reach room temperature before opening.

2 To seal opened pouch, fold end over and apply adhesive tape.

3 Place 3M Petrifilm Coliform Count Plate on

level

surface. Lift top film. With 3M ™ Electronic Pipettor or equivalent held

perpendicular

of bottom film.

to plate, place 1mL of sample or diluted sample onto center 4

Roll

top film down onto sample avoid entrapping air bubbles.

gently Do not

to prevent pushing sample off film and to let top film drop.

5 With

flat

side down, place 3M ™ Petrifilm ™ Spreader on top film over inoculum.

6

Gently Do not

apply pressure on 3M Petrifilm Spreader to distribute inoculum over circular area before gel is formed. twist or slide the spreader. Lift 3M Petrifilm Spreader. Wait a minimum of 1 minute for gel to solidify.

Incubation Interpretation

7

Requester:

Incubate plates with clear side up in stacks of up to 20. It may be necessary to humidify incubator to minimize moisture loss. Susan Barker

See product instructions Creator:

deZinnia_21849

File Name:

Petrifilm_21849-07_FS.ai

Structure #:

Illustration

Date:

04/17/17

Scale:

1 Inch

This artwork has been created as requested by 3M. 3M is responsible for the artwork AS APPROVED and assumes full responsibility for its correctness.

8 3M Petrifilm Coliform Count Plates can be counted using the 3M ™ Petrifilm Plate Reader, on a standard colony counter or other illuminated magnifier. Colonies may be isolated for further identification. Lift top film and pick the colony from the gel.

Use Appropriate Sterile Diluents

Butterfield's phosphate buffered dilution water, 0.1% peptone water, peptone salt diluents, buffered peptone water, saline solution (0.85–0.90%), bisulfite-free letheen broth or distilled water.

For optimal growth and recovery of the microorganisms, adjust the pH of the sample suspension to 6.6-7.2.

Do not use diluents containing citrate, bisulfite or thiosulfate with the 3M Petrifilm Coliform Count Plates; they can inhibit growth.

If citrate buffer is indicated in the standard procedure, substitute with one of the buffers listed above, warmed to 40–45°C.

Coliform Count Plate

5

Bubbles

The illustrations below show examples of various bubble patterns associated with gas producing colonies. All should be enumerated.

3M Food Safety offers a full line of products to accomplish a variety of your microbial testing needs. For more product information, visit us at

3M.com/foodsafety/Petrifilm

or call 1-800-328-6553.

3M Food Safety 3M Center, Building 275-5W-05 St. Paul, MN 55144-1000 USA 1-800-328-6553 3M.com/foodsafety 3M Canada Post Office Box 5757 London, Ontario N6A 4T1 Canada 1-800-364-3577 Coliform Count Plate User’s Responsibilities:

3M Petrifilm Plate performance has not been evaluated with all combinations of microbial flora, incubation conditions and food matrices. It is the user’s responsibility to determine that any test methods and results meet the user’s requirements. Should re-printing of this Interpretation Guide be necessary, user’s print settings may impact picture and color quality.

For detailed CAUTIONS, DISCLAIMER OF WARRANTIES/LIMITED REMEDY and LIMITATION OF 3M LIABILITY, STORAGE AND DISPOSAL information and INSTRUCTIONS FOR USE, see Product’s package insert.

3M and Petrifilm are trademarks of 3M. Used under license in Canada. Please recycle. Printed in USA. © 3M 2017. All rights reserved. 70-2008-4573-6 (Rev-0917) 6

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