1.

Settle Plate Sampling

1.1. Introduction

Settle plate sampling is a direct method of assessing the likely number of microorganisms
depositing onto the product or surface in a given time. In settle plate sampling Petri dishes
containing agar medium are opened and exposed for a given period, thus allowing microbe-
bearing particles to deposit onto them. Petri dishes which are 90 mm in diameter (approximate
internal area 64 cm2) are most commonly used. The number of microbe bearing particles
deposited onto the agar surface of the plate over the period of exposure is ascertained by
incubation of the plate and counting the number of microbial colonies, more commonly known
as colony forming units (CFU). The microbial deposition rate may be reported as the number
depositing in a given area per unit time.

1.2. Sample equipment

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1.3. Sample locations

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1.4. Frequency of sampling

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1.5. Method of sampling

1.5.1. Examine the plates for contamination prior to use.

1.5.2. Assemble the plates required and ensure that the correct information is written on the
base of the plate (the part containing the media) with ink or another marker. Do not
mark the lid of the plate, as there is always a possibility of lids coming off and being
replaced on the incorrect sample plate. The following details must be marked on each
plate:
1.5.2.1. Date and time of day sample was taken;
1.5.2.2. Position/sample number.
1.5.3. Transfer the plates into the area where they are to be exposed as outlined in the
appropriate transfer procedure.
1.5.4. Place the plates in the appropriate positions with the lids still on.
1.5.5. Raise lids to expose the surface of the medium and rest the lid next to the respective
plate. Take care not to put fingers on plates. Avoid passing anything over the top of
plates being exposed, where possible.
1.5.6. Leave plates exposed for 1 hour.
1.5.7. After exposure:
1.5.8. Cover the plates with the respective lids.
1.5.9. Swab areas where plates have been exposed with a suitable disinfectant to remove any
trace of media or condensation from the lids, which may contaminate the cleanroom.
1.5.10. Collect all plates exposed and return to microbiology for incubation. Ensure plates are
secured in a suitable container.
1.5.11. Complete and enclose the necessary documentation.

1.6. Incubation conditions

1.6.1. Following testing, samples should be incubated as soon as possible - within 24 hours of
sampling, same day is preferred - and should be held at room temperature with the
medium uppermost until incubated or manipulated.
1.6.2. Samples are to be incubated, inverted, at 30 °C for at least 4 days is suitable for the
growth of bacteria.
1.6.3. If the medium is dropped or touched by an operator then this should be reported, the
sample should be marked accordingly and treated as usual.
1.6.4. Under no circumstances should samples that have been taken be refrigerated.

1.7. Results and reading of samples

1.7.1. After appropriate incubation microbiological contamination should grow into discrete
macroscopic colonies that can be enumerated and the number of discrete colony
forming units (CFU) can be counted on each sample.
1.7.2. Record the number (per unit surface area or per hour) on the appropriate report.
1.7.3. Colony types may be identified if this is considered appropriate.
1.7.4. If CFU are not discrete (coincidence) entities or are Too Numerous To Count (TNTC -
usually greater than 300 CFU per sample or per hour), record the result as TNTC.
1.8. Warning and Action Limits

1.8.1. Recommended Warning Limit is 10 CFU and the recommended Action Limit is 20 CFU.