Active Air Sampling

Active air sampling is intended to provide an index of the number of microorganisms per unit volume of air space in clean rooms. If the clean room is served by a good air-handling system, with integral HEPA filters in place, airborne microbial contamination arises from personnel operating within the clean room.

Most active air sampling should be done when the clean room is operational. However, if a clean room has been nonoperational for a few days (e.g., a long weekend) or a few weeks (e.g., a scheduled shutdown for vacation or maintenance), it is beneficial to start sampling a few days prior to production start-up.

All active air samplers will disrupt air flow to some extent. They should be located carefully, and when they are operated (none work on the continuous sampling principle), they must not counteract protective air flow patterns in significant parts of the clean room.

Active air sampling is the only microbiological environmental monitoring process involving serious capital expenditure. Active air samplers cost several thousands of dollars, insignificant compared with the costs of production autoclaves, aseptic filling machines, etc., but significant enough in terms of the cost of laboratory equipment for there to be lively competition between suppliers.

It is important to understand the distinctions between active air samplers. Figure 2.1 summarizes some of the main characteristics of available active air samplers.

1. The "traditional" active air sampler is the slit-to-agar sampler of the Andersen sampler, Casella sampler, Mattson Garvin sampler, etc. types as shown diagrammatically in Figure 2.1. These have become the "standard" against which other samplers are compared.

Slit-to-agar samplers of this type have been widely used for monitoring pharmaceutical clean rooms and have in fact dominated the market for many years.

Agar Plate

Agar Plate

Pump

Flexible Hose

Figure 2.1. Simplified representation of a slit-to-agar sampler.

They are heavy, thus limiting their portability, even though they are usually mounted on trolleys or carts to provide some flexibility, and they can be fumigated but are difficult to disinfect. They are best dedicated to one clean room and maintained "captive."

The Petri dishes used in slit-to agar samplers are of a nonstandard size (150 mm) requiring a fill volume of about 100 ml. They are unsuited to most automated plate pourers. The cost of media is increased at least five-fold over samplers that use standard size Petri or Rodac Petri dishes.

2. A second type of active sampler is the Reuter Centrifugal Sampler (RCS). Two types of instrument are marketed, the RCS and the RCS-Plus. Both are battery-powered, lightweight, portable and easy to fumigate and disinfect.

In the first development of the RCS (still marketed and used widely), air is drawn into an open-fronted cylindrical housing by means of a low-pitch impeller (Figure 2.2). The air drawn into the housing is redirected back out again through a turn of about 360°. Some of the air is forced towards the inner wall of the housing, where an agar-containing flexible plastic strip is located around the circumference. The cone of air deflected forward from the RCS sampler interferes with protective airflow patterns in areas where it has been used (Kaye, 1988).

In its later development, the RCS-Plus, the air is exhausted through ports at the rear of the impeller head, successfully reducing the air-flow interference effect (Ljungqvist and Reinmuller, 1991).

The earlier of the two developed instruments, the RCS, has a stated air-intake rate of 280 litres per minute of which, the manufacturers claim, about one-seventh (40 litres of air per minute) is directed onto the agar strip. There has been much debate

Strip

Figure 2.2. Simplified representation of an impeller head of an RCS active air sampler.

Strip

Figure 2.2. Simplified representation of an impeller head of an RCS active air sampler.

as to whether this instrument or its successor is capable of collecting all particle sizes that might carry airborne microorganisms, if it even collects particles of the sizes most likely to carry airborne microorganisms, and if it can be regarded as a truly quantitative instrument for measurement of airborne microorganisms against published limits.

It seems that the RCS centrifugal sampler is less efficient at capturing airborne microorganisms than other types of available samplers. The question though is: Does it really matter?

The answer: It depends on the circumstances. The RCS effectively samples at 40 litres per minute for a maximum sampling time of five minutes, to a maximum sample volume of 400 litres. It is unsuited to sampling E.U. Grade A environments (laminar flow-protected aseptic areas) because, even if there were no controversy over other aspects of this sampler, it quite simply does not draw a large enough sample to verify compliance with the limit of less than 1 cfu per m3 (1000 litres).

Its limitations are insignificant in Grade C environments against a limit of no more than 100 cfu per m3 (preparation areas for aseptic manufacture or filling rooms for terminally sterilized products), but borderline for Grade B environments (aseptic filling rooms). It is probably wisest to confine this instrument to Grade C environments for sterile manufacture, and to sampling nonsterile manufacturing environments.

The RCS-Plus instrument samples at 50 litres per minute and can be operated for 20 minutes to collect a 1-m3 air sample. There are data demonstrating that its efficiency of collection of microorganisms on particles of 4 ^m with which most microorganisms are believed to be associated, and larger, is comparable to slit-to-agar samplers (Benbough, 1992). This instrument is suitable for sampling all grades of pharmaceutical clean room, is less robust than the RCS, and requires frequent calibration of its sampling heads.

Centrifugal samplers are generally in the same price range as other types of samplers but running costs are higher because of the need to purchase the agar-containing flexible strips. Empty strips can be purchased for local laboratory filling but involve introducing a nonstandard operation into laboratories.

3. Filtration is the third means of active air sampling, but death of microorganisms by desiccation on the membranes has restricted its application. The most widely used variant on filtration, the Sartorius MD-8 sampler, avoids desiccation by using gelatin membranes. After sampling, the gelatin membrane is transferred to an agar plate where it dissolves and merges into the agar during incubation (plates should be well dried before transfer to prevent colonies from spreading). Efficiencies of collection and recovery of microorganisms are comparable to slit-to-agar samplers (Parks et al., 1996; Pendlebury and Pickard, 1997).

The MD-8 is battery-powered, heavier and bulkier (because of the pump system) than the centrifugal samplers but is still portable and easy to fumigate and disinfect.

The maximum sampling speed of the MD-8 is about 130 litres per minute. Flow rates are adjustable to match the local airflow and enable isokinetic sampling. This makes the MD-8 the most useful of all the samplers for evaluating the airborne microbial counts in laminar flow-protected areas (e.g., at point-of-fill) while operational. All other active air samplers are required to be used with considerable care in these areas, to avoid the risk of disrupting protective air flow. This is greater than the benefit of having data to show that microorganisms have been excluded.

4. At least three variants of another type of air sampler exist, less commonly used than in monitoring pharmaceutical clean rooms. These operate on the principle that air is drawn through a perforated atrium head where air samples are collected by impaction on an agar plate. The air is exhausted at the rear (beneath the agar plate). The location of the exhaust forces the sampled air, after its initial impaction, to turn through 90° and flow over the surface of the agar to exhaust. This may help to improve microbiological collection efficiency.

The heaviest of the three models is comparable to the MD-8 with respect to portability and ease of fumigation and disinfection. Models are marketed by PBI International, Merck and by Veltek Associates (VAI) (Figure 2.3).

These samplers are operated by constant-speed pumps, and sample volumes are controllable through a timer setting.

The agar used in the Merck and VAI samplers is poured in standard 90-mm Petri dishes and for the PBI sampler in Rodac (55-mm) Petri dishes. Their running costs are far less than those of the other sampler types.

Figure 2.3. Simplified representation of an active air sampler with perforated atrium head.

Atrium Top

Agar Plate

Figure 2.3. Simplified representation of an active air sampler with perforated atrium head.

Table 2.2 summarizes some of the characteristics of available active air samplers.

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