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Determination of Enterobacteriaceae
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5) Incubator, 35-37°C.
6) Colony counter (Quebec dark field model or equivalent recommended).
7) Tally register .
8) Violet-Red Bile Glucose Agar to be pretested for productivity and selectivity.
9) Glucose Salt Medium.
10) Nutrient Agar.
11) Tetramethylparaphenylenediamine dihydrochloride, 1% aqueous solution.
12) Filter paper, Whatman No. 2, pieces 6 cm square.
13) Colworth Stomacher 400, blender or equivalent.
7. Procedure
Analyze each sample unit individually.
The test shall be carried out in accordance with the following instructions:
7.1 Handling of Sample Units
7.1.1 In the laboratory prior to analysis, except for shelf-stable foods, keep sample units refrigerated (0-5°C) or frozen depending on the nature of the product. Thaw frozen samples in a refirgerator, or under time and temperature conditions which prevent microbial growth or death.
7.1.2 Analyze sample units as soon as possible after receipt in the laboratory.
7.2 Preparation of Media
7.2.1 Prepare media for either the Plating Method (7.4) or MPN procedure (8) and dispense in appropriate quantities. Sterilize.
7.2.2 Clean surface of working area with a suitable disinfectant.
7.3 Preparation of Sample
7.3.1 Have ready 0.1% peptone water diluent, and tempered VRBG agar (7.4.2).
7.3.2 To ensure a truly representative analytical portion agitate liquid or free flowing materials until the contents are homogeneous. If the sample unit is a solid, obtain the analytical unit by taking a portion from several locations within the sample unit.
7.3.3 Prepare a 1:10 dilution of the food by aseptically adding 11(10) g or mL (the analytical unit) into 99(90) mL of the diluent. Blend or shake according to the type of food as indicated in Table I.
Note: Volume in Brackets indicates alternate procedure for making dilutions.
7.3.3.1 If a homogeneous suspension is to be obtained by blending, the blending time should not exceed 2.0 min in order to prevent over-heating. With foods that tend to foam, use blender at low speed, and remove aliquot from below liquid/foam interface.
7.3.3.2 If a homogeneous suspension is to be obtained by shaking, shake the dilution bottles 25 times through a 30 cm arc in approximately 7 sec.
