Environmental monitoring: settle plates
One of the series of learning articles, an overview of settle plates for environmental monitoring.
Settle plates are Petri-dishes, typically of either 9cm or 14cm diameter, containing different fill volumes of agar (normally between 20 and 30 mL). Settle plates are designed to detect any viable micro-organisms that may directly settle on or in the product (that is micro-organisms that are carried in the air-stream, although a person who leans over a plate can also potentially deposit micro-organisms). At determined monitoring locations (ideally positioned and exposed either side of the testing environment) the lids of the dishes are removed and the plates are exposed to the air for a defined period of time. In theory, micro-organisms and units containing micro-organisms settle out of the air under gravity, and are deposited onto horizontally positioned agar plates. This theoretically works better in turbulent or laminar airflows. The efficiency can be described as the ‘settling rate’.
The settling rate depends partly on the characteristics of the particles and on the air-flows. Larger units will tend to settle faster (due to gravitational effects) and settling is facilitated by still air-flows (which should not occur within a correctly designed uni-directional air-flow zone). Smaller particles have a lower tendency to settle due to sir resistance and air currents. The principle behind settle plates is that most micro-organisms in air are in association with particles. Generally the ‘complete particle’ (micro-organism in association with the ‘carrier’) is 12mm diameter or larger[i].
Outside of uni-directional air, such as the main cleanroom itself, then the greater the degree of turbulence there is. The amount of air turbulence is proportional to the amount of time that particles remain suspended in the air. Thereby, the greater the amount of air turbulence then the longer the particles will remain suspended in the air (this is not always a bad thing, as particles can be blown away from a critical zone, depending upon the design of the room). This can, however, influence the reliability of the settle plate and here the additional use of active air-samplers can provide additional assurance for the microbiologist assessing the cleanroom cleanliness.
The phenomenon of gravitational settling is, however, a debatable issue. The prevailing view, as discussed above, is that as most micro-organisms are associated with physical particles they will be large enough to settle out of the air due to gravity i. The dissenting view is that micro-organism carrying particles or any micro-organisms not associated with units as being light enough to remain in the air-stream for several minutes and possibly be carried out of the air-stream and not settle[ii]. Much of this debate thereby centres on the size of the particles in the air and the airflow.
The exposure time of the settle plate can be varied, although there is probably little value in exposing plates for less than one hour. For consistency of sampling, for aseptic filling, the EU GMP Guide recommends a four hour exposure time. This time should not be exceeded without strong justification, and even then there will probably be a challenge from the regulatory authority. For exposure times under four hours, such as when a shorter activity is being monitored, the result obtained should be extrapolated using the simple equation:
Count x 240 = cfu / 4 hours
Time exposed (minutes)
The risk from any exposure is desiccation. The depth and condition of the agar are the key variables, as is the cleanroom environment. The agar in the plate will dry out faster if the airflow is excessively high or if the air humidity is low. Therefore the exposure time of settle plates under the conditions of use (a particular cleanroom or uni-directional airflow cabinet) must be validated.
[i] Whyte, W. (1986): ‘Sterility assurance and models for assessing airborne bacterial contamination’, Journal of Parenteral Science and Technology, 40, pp188-197
[ii] Sykes, G. (1970): ‘The control of airborne contamination in sterile areas’, Aerobiology: Proceedings of the 3rd International Symposium, in Silver, I. H. (ed.), Academic Press, London
REF: http://pharmig.blogspot.com/2010/07/environmental-monitoring-settle-plates.html ACESSADO EM 08/03/12
quinta-feira, 8 de março de 2012
Environmental monitoring: settle plates
One of the series of learning articles, an overview of settle plates for environmental monitoring.
Settle plates are Petri-dishes, typically of either 9cm or 14cm diameter, containing different fill volumes of agar (normally between 20 and 30 mL). Settle plates are designed to detect any viable micro-organisms that may directly settle on or in the product (that is micro-organisms that are carried in the air-stream, although a person who leans over a plate can also potentially deposit micro-organisms). At determined monitoring locations (ideally positioned and exposed either side of the testing environment) the lids of the dishes are removed and the plates are exposed to the air for a defined period of time. In theory, micro-organisms and units containing micro-organisms settle out of the air under gravity, and are deposited onto horizontally positioned agar plates. This theoretically works better in turbulent or laminar airflows. The efficiency can be described as the ‘settling rate’.
The settling rate depends partly on the characteristics of the particles and on the air-flows. Larger units will tend to settle faster (due to gravitational effects) and settling is facilitated by still air-flows (which should not occur within a correctly designed uni-directional air-flow zone). Smaller particles have a lower tendency to settle due to sir resistance and air currents. The principle behind settle plates is that most micro-organisms in air are in association with particles. Generally the ‘complete particle’ (micro-organism in association with the ‘carrier’) is 12mm diameter or larger[i].
Outside of uni-directional air, such as the main cleanroom itself, then the greater the degree of turbulence there is. The amount of air turbulence is proportional to the amount of time that particles remain suspended in the air. Thereby, the greater the amount of air turbulence then the longer the particles will remain suspended in the air (this is not always a bad thing, as particles can be blown away from a critical zone, depending upon the design of the room). This can, however, influence the reliability of the settle plate and here the additional use of active air-samplers can provide additional assurance for the microbiologist assessing the cleanroom cleanliness.
The phenomenon of gravitational settling is, however, a debatable issue. The prevailing view, as discussed above, is that as most micro-organisms are associated with physical particles they will be large enough to settle out of the air due to gravity i. The dissenting view is that micro-organism carrying particles or any micro-organisms not associated with units as being light enough to remain in the air-stream for several minutes and possibly be carried out of the air-stream and not settle[ii]. Much of this debate thereby centres on the size of the particles in the air and the airflow.
The exposure time of the settle plate can be varied, although there is probably little value in exposing plates for less than one hour. For consistency of sampling, for aseptic filling, the EU GMP Guide recommends a four hour exposure time. This time should not be exceeded without strong justification, and even then there will probably be a challenge from the regulatory authority. For exposure times under four hours, such as when a shorter activity is being monitored, the result obtained should be extrapolated using the simple equation:
Count x 240 = cfu / 4 hours
Time exposed (minutes)
The risk from any exposure is desiccation. The depth and condition of the agar are the key variables, as is the cleanroom environment. The agar in the plate will dry out faster if the airflow is excessively high or if the air humidity is low. Therefore the exposure time of settle plates under the conditions of use (a particular cleanroom or uni-directional airflow cabinet) must be validated.
[i] Whyte, W. (1986): ‘Sterility assurance and models for assessing airborne bacterial contamination’, Journal of Parenteral Science and Technology, 40, pp188-197
[ii] Sykes, G. (1970): ‘The control of airborne contamination in sterile areas’, Aerobiology: Proceedings of the 3rd International Symposium, in Silver, I. H. (ed.), Academic Press, London
REF: http://pharmig.blogspot.com/2010/07/environmental-monitoring-settle-plates.html ACESSADO EM 08/03/12
One of the series of learning articles, an overview of settle plates for environmental monitoring.
Settle plates are Petri-dishes, typically of either 9cm or 14cm diameter, containing different fill volumes of agar (normally between 20 and 30 mL). Settle plates are designed to detect any viable micro-organisms that may directly settle on or in the product (that is micro-organisms that are carried in the air-stream, although a person who leans over a plate can also potentially deposit micro-organisms). At determined monitoring locations (ideally positioned and exposed either side of the testing environment) the lids of the dishes are removed and the plates are exposed to the air for a defined period of time. In theory, micro-organisms and units containing micro-organisms settle out of the air under gravity, and are deposited onto horizontally positioned agar plates. This theoretically works better in turbulent or laminar airflows. The efficiency can be described as the ‘settling rate’.
The settling rate depends partly on the characteristics of the particles and on the air-flows. Larger units will tend to settle faster (due to gravitational effects) and settling is facilitated by still air-flows (which should not occur within a correctly designed uni-directional air-flow zone). Smaller particles have a lower tendency to settle due to sir resistance and air currents. The principle behind settle plates is that most micro-organisms in air are in association with particles. Generally the ‘complete particle’ (micro-organism in association with the ‘carrier’) is 12mm diameter or larger[i].
Outside of uni-directional air, such as the main cleanroom itself, then the greater the degree of turbulence there is. The amount of air turbulence is proportional to the amount of time that particles remain suspended in the air. Thereby, the greater the amount of air turbulence then the longer the particles will remain suspended in the air (this is not always a bad thing, as particles can be blown away from a critical zone, depending upon the design of the room). This can, however, influence the reliability of the settle plate and here the additional use of active air-samplers can provide additional assurance for the microbiologist assessing the cleanroom cleanliness.
The phenomenon of gravitational settling is, however, a debatable issue. The prevailing view, as discussed above, is that as most micro-organisms are associated with physical particles they will be large enough to settle out of the air due to gravity i. The dissenting view is that micro-organism carrying particles or any micro-organisms not associated with units as being light enough to remain in the air-stream for several minutes and possibly be carried out of the air-stream and not settle[ii]. Much of this debate thereby centres on the size of the particles in the air and the airflow.
The exposure time of the settle plate can be varied, although there is probably little value in exposing plates for less than one hour. For consistency of sampling, for aseptic filling, the EU GMP Guide recommends a four hour exposure time. This time should not be exceeded without strong justification, and even then there will probably be a challenge from the regulatory authority. For exposure times under four hours, such as when a shorter activity is being monitored, the result obtained should be extrapolated using the simple equation:
Count x 240 = cfu / 4 hours
Time exposed (minutes)
The risk from any exposure is desiccation. The depth and condition of the agar are the key variables, as is the cleanroom environment. The agar in the plate will dry out faster if the airflow is excessively high or if the air humidity is low. Therefore the exposure time of settle plates under the conditions of use (a particular cleanroom or uni-directional airflow cabinet) must be validated.
[i] Whyte, W. (1986): ‘Sterility assurance and models for assessing airborne bacterial contamination’, Journal of Parenteral Science and Technology, 40, pp188-197
[ii] Sykes, G. (1970): ‘The control of airborne contamination in sterile areas’, Aerobiology: Proceedings of the 3rd International Symposium, in Silver, I. H. (ed.), Academic Press, London
REF: http://pharmig.blogspot.com/2010/07/environmental-monitoring-settle-plates.html ACESSADO EM 08/03/12
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