Cleanroom Monitoring Basics

Cleanroom classifications as specified by different standards are shown below in Table 1.

Sterile products are required to be filled and stoppered in an ISO Class 5 or EU GMP Class A environment.  Non-sterile products are typically produced in an ISO Class 8 or EU GMP Class D environment. 

Most Biopharmaceutical Quality Managers create their risk assessment using a conflation of the most conservative standard requirements (shown in Table 1).

TABLE 1.

Obsolete
Fed Std 209E
ISO
14644-1
EU GMP,
Annex 1
Max. Concentration Limits
(particles/m3 of air)
for particles equal to and larger than
the sizes listed below
0.3μm 0.5μm(1) 1μm 5μm (2)
  ISO Class 4.8 Grade A not defined 3,520 not defined 20
100 ISO Class 5 Grade B 10,200 3,520 832 29
1,000 ISO Class 6 - - - 102,000 35,200 8,320 290
10,000 ISO Class 7 Grade C not defined 352,000 83,200 2,900
100,000 ISO Class 8 Grade D not defined 3,520,000 832,000 29,000

(1) EU GMP Annex 1 and ISO 14644-1:2015 limits are identical on the 0.5 μm particle size "at rest". 

(2) EU GMP, Annex 1 requirements for 5 µm. Virtually every pharmaceutical monitors 5µm particles in Class 5 areas as a key indicator of biocontamination risk assessment.

TABLE 2.

Max. Microbiological Active Air Action Level
Colony Forming Units (cfu) per cubic meter (average values)
ISO 14644-1 Class
(GMP Grade)
FDA Pharmaceutical CGMP:2004 EU GMP, Annex 1 WHO, Tech Report Series, No. 902, 2002
4.8 (A) Not Defined <1 <3
5 (B) 1 10 10
6 7 Not Defined Not Defined
7 (C) 10 100 100
8 (D) 100 200 200

With the release of EU GMP:2008 and ISO 14644, and with the increase importance of 5 micron particle to the Life Science Industry, cubic meter sampling became a requirement for ISO Class 5 (EU Class A) environments.

Sample Times

When sampling a one cubic meter of air in an ISO Class 3-5 Cleanroom, the sample time is a function of the particle counter's flow rate, as GMP requires a cubic meter sample for Grade A/B zones (regulatory minimum).  As a best practice, the life science industry typically use full cubic meter sampling for ISO Class 5.

0.1 CFM equals 2.83 LPM flow rate, and takes 5 hours 53 minutes per sample.
1 CFM equals 28.3 LPM flow rate, and takes 35.3 minutes per sample.
50 LPM flow rate takes 20 minutes per sample.
75 LPM flow rate takes 13.3 minutes per sample.
100 LPM flow rate takes 10 minutes per sample. (recommended with spot or sequential monitoring)

** Fill-Stations or other continuous monitoring  applications (aka "Critical Areas") should use a 1 CFM particle counter.

When sampling air in an ISO Class 6 cleanroom:

0.1 CFM equals 2.83 LPM flow rate, and takes 24 minutes 6 seconds per sample.
1 CFM equals 28.3 LPM flow rate, and takes 2 minutes 25 seconds per sample.
50 LPM flow rate takes 1 minute and 22 seconds per sample.
75 LPM flow rate takes 1 minute per sample (regulatory minimum)
100 LPM flow rate takes 1 minute per sample (regulatory minimum)

As you can see, if you have an ISO Class 5 cleanroom, labor efficiencies can be attained by using higher flow rate particle counters when 5 μm is a size of interest (e.g., life science industry).  The additional cost for a higher flow rate instrument can generally be recovered from less labor hours required to do sampling/monitoring.  If doing sequential monitoring, you can use the the same high flow rate particle counter in higher ISO/GMP classifications.

 

Good Monitoring Techniques

There is a difference between cleanroom monitoring and certification/validation. 

The number of sample locations when certifying or validating a cleanroom is determined by a chart provided in the ISO 14644 standard.

However, when doing day-to-day monitoring of the room, you are only trying to verify that the air enveloping the process is adequately clean enough to "well exceed" regulatory standards. Processes which generate submicron particles include:

Process and machinery. Basically, anything that creates friction is creating particles. 

People: skin flakes and oil, cosmetics and perfume, spittle, clothing debris (lint, fibers, etc.), and hair.  People are a major sources of particles.

Fluids:  Particles floating in air, bacteria, organics and moisture, floor finishes or coatings, cleaning chemicals, plasticizers (outgasses), and deionized water.

Simply, any kind of process, machinery, and especially people add to the particle burden of the room.

Best Practices

Zero Count Test (also called the Purge Test): Before you begin your daily round of sampling, it is good practice to install a purge filter onto the inlet, and perform a one-minute False Count Test (also known as a Zero Count Test) to ensure the particle counter does not have a fundamental or catastrophic failure.  More Information Here

Sample Averaging: In a particular location, take three samples, then average the counts. If you begin monitoring immediately, and you take (for example) three one minute samples to average you will likely see that the 5 micron count on each sample decreases while the 0.5 micron count does not change significantly. This is because 5 micron particles fall out more quickly than 0.5 micron particles. 

Air Settlement: 
When monitoring a cleanroom it is good practice to let the air settle prior to taking a sample.  Many of our customers program a minute or two delay into their particle counter so that the sampling begins after the air in the sample area is allowed to settle.  Again, this allows the person taking the sample an opportunity to at least step back 6-10 feet to allow the air to settle (remember, people are a source of particles).  You are also monitoring a process, and therefore evacuating an entire cleanroom to do monitoring is not recommended. 

Sample Height:  
It is good practice to take the sample from a height of about one meter or "at work height" above the floor.  This would be from a counter top or often from a cart.  With regards to monitoring filters, unless you are doing a filter scan you are not likely to pick-up any leaks... simply, it is a wasted effort.

Trend Analysis:  
Particle counts should be uploaded into a database to allow for trend analysis.  The whole idea of trend analysis is to identify worsening trends before it becomes a deviation, which requires a considerable amount of paperwork.  By identifying an early trend, you can start your investigation and take early corrective actions before a problem occurs.  Increasing counts may occur slowly over the course of months, and those performing the monitoring should know the normal particle burden of the cleanroom as well as any trends. There are certainly LIMS solutions that large pharmaceuticals utilize. For the smaller biotech or pharmaceutical firms, Climet offers a proprietary software solution that's a fraction the cost of a LIMS system. 

Sample Location: 
Generally, you want to sample in close proximity to the process, so that you are sampling the air that envelopes the process; but not too close to a process that inherently generates particles.  Being too close may skew the sample data, and may contaminate the particle counter sensor.  This can be especially tricky when you are involved in a fill-process, where a powder or liquid spray may be sucked into the sample probe.  For these applications, testing the position of the probe should be part of the validation process.  Please note, 1 CFM particle counters are best suited for fill-processes or continuous monitoring applications. Fill-processes require continuous monitoring (before, during and after). High flow rates may have an adverse impact as they pull air in from a wider radius. 

Microbial Monitoring in the Life Science Industry (Sterile Environments):
In Life Science industries these organizations should be measuring both particle concentration, and for sterility.  The particle concentration is monitored with a particle counter; and sterility is monitored through a microbial air sampler.  The particle counter's function is to provide early warning to a serious event in order to allow users to stop the process before expensive materials are wasted.  The microbial sampler demonstrates that the area was sterile and that product can be released.  According to EU GMP (Annex 1, #18, #19), "Where aseptic operations are performed monitoring should be frequent."  Microbial sampling must be part of your normal monitoring protocol.

Conservative Interpretation
It is good practice to conservatively interpret the ISO and EU GMP standards, and to always put quality above price. Failing to follow this advise not only puts expensive production at risk, but those who self-interpret may also risk heavy fines levied by the FDA or GMP, and even plant closures.  It is recommended that terms in the standards such as 'should' or 'recommended' be interpreted as 'shall' and 'must'

 


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Climet Instruments Company
A Division of Venturedyne, Ltd.

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