6.1.2 Analyze samples as soon as possible after their receipt in the laboratory.

6.2.1 Have sterile diluent ready. Disinfect the surface of the work area.

6.2.2 Place the Petrifilm plate on a flat surface. Mark identifying sample information on the film.

6.3.1 To ensure a representative analytical unit, agitate liquids until the contents are homogeneous. If the food sample is a solid, take representative portions from several locations within the sample. Prepare a 1:10 dilution in an appropriate (see 5.2) diluent. Mix thoroughly.

6.3.2 For acid products adjust pH of the diluted sample to 6.6 - 7.2 with 1N NaOH.

6.4.1 Lift the top film and carefully inoculate 1 mL of sample or diluted sample in the center of bottom film. Either a pipette or a pipettor can be used for sample addition.

6.4.2 Replace top film. Carefully roll top film down to avoid entrapping air bubbles.

6.4.3 Distribute sample evenly using a downward pressure on the center of the plastic spreader, flat side down. Do not slide the spreader across the film. Leave plate undisturbed for at least 1 minute to permit the gel to solidify.

6.4.4 Return unused plates to foil pouch. Seal pouch by folding and taping the open end. Store resealed foil pouch in a cool dry place. Use plates within one month after opening pouch. Exposure of Petrifilm plates to temperatures above 25^oC and/or humidities >50% RH can affect the performance of the plates. Do not use plates that show orange or brown discoloration. Expiration date and lot number are Noted on each package of Petrifilm plates. The lot number is also Noted on individual test films.

6.4.5 Incubate plates in a horizontal position with the clear side up in stacks not exceeding 20 units. Follow current industry standards for incubation temperature. Incubate plates 24 ± 2 hours. Examine for E. coli and other coliforms. Some E. coli colonies require additional time to form the blue precipitate. Reincubate plates an additional 24 ± 2 hours to detect any additional E. coli (7.1 and 7.2).

6.5.1 Count plates promptly after incubation period. If impossible to count at once, store plates in the freezer. This should be avoided as a routine practice.

6.5.2 Use a standard colony counter for counting purposes. A magnified-illuminator may also be used to facilitate counting.

6.5.3 The circular growth area is approximately 20 cm². Estimates can be made on plates containing more than 150 colonies by counting the average number of colonies in one or more representative squares and determining the average number per square. Multiply the average number by 20 to determine total count per plate.