1. ISO 14644-1:2015
CLEANROOMS & ASSOCIATED CONTROLLED
ENVIRONMENTS
PART 1: CLASSIFICATION OF AIR CLEANLINESS
BY PARTICLE CONCENTRATION
Anwar Munjewar
Quality Assurance Department
Bharat Serums and Vaccines Limited

2. ABOUT INTERNATIONAL
ORGANIZATION FOR
STANDARDIZATION■ ISO is an independent, non-governmental international organization with a
membership of 163 national standards bodies. Through its members, it brings
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■ ISO has published more than 21000 International Standards and related
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agriculture and healthcare. ISO International Standards impact everyone,
everywhere.

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STANDARDS?
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Final draft
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members
ISO
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If proposal is accepted
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vote

4. ISO TECHNICAL COMMITTEE 209
Cleanroom associated with controlled environments
■ Participating Country :
22
■ Observing Country : 19
Secretariat: USA (ANSI)
Australia (SA) Kenya (KEBS)
Belgium (NBN) South Korea (KAT)
Brazil (ABNT) Netherlands (NEN)
China (SAC) Norway (SN)
Denmark (DS) Phillipense (BPS)
Finland (SFS) Portugal (IPQ)
France (AFNOR) Russian Federation
Germany (DIN) Sweden (SIS)
Ireland (NSAI) Switzerland (SNV)
Italy (UNI)
United Kingdom
(BSI
Argentina Poland
Bosnia and Herzegovina Romania
Bulgaria Saudi Arabia
Cuba Serbia
Czeh Republic South Affrica
Egypt Thailand
Hungary Turkey
India Ukraine
Iran
Jamaica
Malaysia

5. ISO PARTS & STATUS
■ Part 1: Classification of air cleanliness by particle concentration (15th Dec 2015)
■ Part 2: Monitoring to provide evidence of cleanroom performance related to air
cleanliness by particle concentration (15th Dec 2015)
■ Part 3: Test Methods (DIS 2014)
■ Part 4: Design, construction and start-up (2001)
■ Part 5: Operations (2004)
■ Part 6: Vocabulary (2006)
■ Part 7: Separative devices (clean air hoods, gloveboxes, isolators and environments)
(2007)
■ Part 8: Classification of air cleanliness by Chemical concentration (2013)
■ Part 9: Classification of surface cleanliness by Particle concentration (2012)
■ Part 10: Classification of surface cleanliness by Chemical concentration (2013)
Note: There is no Part 11 in ISO 14664, and Part 12 – Part 16 is under process(Draft).

6. SCOPE
■ Only particle populations having cumulative distributions based on threshold
(lower limit) particle sizes ranging from 0.1 µm to 5.0 µm are considered for
classification purposes.
■ Light scattering (discreet) airborne particle counters (LSAPC) is the basis
for the determination of the concentration of airborne particles equal to and
greater than the specified sizes, at designated sampling locations.
■ ISO 14644 does not provide for the classification of particle populations that are
outside the specified lower threshold particle size range 0.1 µm to 5.0 µm.
■ An M descriptor may be used to quantify populations of macroparticles
(particles larger than 5 µm).
■ The standards also allow the addition of location to the minimum number of
sampling locations.
■ Calculation for Number of location changed and 95% UCL completely
removed.

7. ISO Class Number

8. Method for Classification of Air
Cleanliness by Particle Concentration
■ Apparatus: Light Scattering Airborne Particle Counter (LSAPC)
■ Establishment of Sampling locations:
– Drive the number of sampling locations from the Table 1
– Positioning the sampling locations
– Sampling locations for large cleanrooms or clean zone
– Establishment of single sample volume and sampling time per locations
– Processing of results
– Interpretation of result

9. A.4.1 Sampling locations related to cleanroom area
AREA LOCATIONS AREA LOCATIONS
2 1 76 15
4 2 104 16
6 3 108 17
8 4 116 18
10 5 148 19
24 6 156 20
28 7 192 21
32 8 232 22
36 9 276 23
52 10 352 24
56 11 436 25
64 12 636 26
68 13 1000 27
72 14 >1000 Formula

10. A.4.2 Positioning the sampling locations
 Use the minimum number of sampling locations NL derived from the Table 1
 Divide the whole cleanroom or clean zone into NL section of equal area
 Select within each section a sampling location considered to be
representative of the characteristics of the section
 Additional sampling locations may be selected for locations considered
critical
A.4.3 Sampling locations for large cleanrooms or clean zone
 When the area of a clean room is greater than 1000 meter square.
 𝑁 𝐿 = 27 𝑥
𝐴
1000
 NL- is the minimum number of sampling locations to be evaluated
 A- is the area of the clean room in meter square.

11. A.4.4 Establishment of single sample volume and sampling
time per locations
 At sample location, sample a volume of air sufficient to detect a minimum of 20
particles if the particle concentration for the largest selected particle size were at the
class limit for the designated ISO class
 Single sample volume:
𝑉𝑠 = (
20
𝐶𝑛,𝑚
) x 1000
Where,
Vs – is the minimum single volume per location (expressed in liters)
Cn,m – is the class limit (number of particles per cubic meter) for the largest
considered particle size.
20 – is the number of particles that could be counted if the particle concentration
were at the class limit.

12. A.6 Processing of results:
A.6.1 Recording of results:
– Record each sample measurements as the number of particles in each single
sample volume.
– When two or more single sample volume are taken at a location, calculate and
record the average number of particles per location at each considered particle
size.
– For particle counters with concentration calculation mode, the manual
evaluation may not be necessary. (i.e. counts per litter to counts per cubic
meter)
A.6.2 Interpretation of result:
– If an out-of-specification count found at a location due to an identified abnormal
occurrence, then that count can be discarded and noted as such on the test
report and a new sample taken.
– If an out-of-specification count found at a location is attributed to a technical
failure of the cleanroom or equipment, then cause should be identified, remedial
action taken and resting performed of the failed sampling location, the
immediate surroundings locations and any other locations affected. The choice
shall be clearly documented and justified.

13. ISO 14644-1:2015(E) ANNEX
■ Annex A – Reference Method For Classification Of Air Cleanliness By Particle
Concentration
■ Annex B – Examples Of Classification Calculations
■ Annex C – Counting And Sizing Of Airborne Macroparticles (M descriptor)
■ Annex D – Sequential Sampling Procedure : NA
■ Annex E – Specification Of Intermediate Decimal Cleanliness Classes And
Particle Size Thresholds : NA
■ Annex F – Test Instruments : NA

14. Why Does the Pharmaceutical Industry
Love 5.0 μm particles?
■ EU inspectors maintain that large particles are potential carriers (hitch-hikers) of, or
are viable organisms themselves. If these particles are present in an aseptic
environment, they represent an increased risk of contamination of the sterile
product. Most ISO Class 5 (Grade A) clean zone shave counts of zero or one on the
>= 5 μm size channel. Viable microorganisms, whose individual sizes are generally
less than 1 μm, tend to form in pairs, chains, and clusters. These colony forming
units together often have a size greater than 5 μm. Therefore, a particle counter is
like the “canary in a coal mine”, because it provides an early warning of a potential
problem.
■ It is worth while noting that USFDA cGMP 2004 does not require measurement of
5um particles in ISO Class 5.

15. Counting and Sizing of Airborne
Macroparticles – M Descriptor
■ The test method describe the measurement of airborne particles with a threshold
size ≥ 5 µm in diameter(macroparticles).
■ The number of sampling location, location selection and quantity of data required
should be in accordance with A.4.
■ M descriptor format:
“ISO M (a;b); C”
Where,
a- is the maximum permitted concertation of macroparticles (in cubic meter)
b- is the equivalent diameter associated with the specified method for
measuring macroparticles.
c- is the specified measurement method.
Example : To express an airborne concentration of 29 particles per cubic meter in the
particle size range ≥ 5 µm using LSAPC
“ ISO M (29; ≥ 5 µm ); LSAPC”

16. Annex A of ISO 14644-1:2015
■ Clause A.2 Apparatus Requirements, under A.2.2 Instrument Calibration
specifies that the particle counter shall have a valid calibration certificate, the
frequency and method of calibration should be based upon ISO 21501-4
■ Old ISO14644 specified calibration be performed by suitable method. New
revision recognises some of the older particle counters (ex. those manufactured
prior to 2011) cannot be calibrated to all of the required tests in ISO21501-4
■ Some details of this new calibration standard are given in the next two slides, a
detailed presentation is available from the author.

17. ISO 21501-4 Calibration
■ Parameters that will now be required
– Size Calibration (Using NIST traceable particles)
– Size Verification
– Counting Efficiency (50% and 100%)
– Size resolution (equal to or less than 15%)
– False Count Rate (1 or less in 15 minutes)
– Maximum Concentration (before coincidence loss)
– Flow Rate (Uncertainty of 5% or less)
– Sample Time (Uncertainty of 1% or less)
– Response Rate (0.5% or less)
– Calibration Interval ( not to exceed 1 year)
– Test Report (minimum information required)

18. Summary
ISO14644-1:1999 ISO14644-1:2015
Title Change
Part 1 : Classification of air cleanliness
Part 1 : Classification of air cleanliness by particle
concentration
Ultra and Macro (U,M) Descriptors
Smaller than 0.1 microns defined as U, >than 5
microns defined as M.
Smaller than 0.1 micron particles will no longer
exist in ISO 14644-1. This issue addressed in
ISO 14644-12 (Air cleanliness by nanoparticles).
Annex A Informative (Graphical Illustration of
classes)
Annex A (normative) Reference method for
determination of classification
Table E.1 of Annex E provides permitted
intermediate air cleanliness classes in increments
of 0.5 (3.5, 4.5, 5.5etc.)
No graphical illustration

19. Summary
ISO14644-1:1999 ISO14644-1:2015
ISO Class 5, 5.0 micron – 29 Particles No more Class 5, 5.0 micron size
Sample Numbers and Locations
Square-root method for sample numbers.
Grid for sample locations
Table A.1. (Fixed numbers). Sections of equal
area, within each section select location
representative of characteristics of the section
Apparatus for Particle Count
DPC = Discrete Particle Counter
LSAPC = Light Scattering Airborne Particle
Counter
ParticleSizes in Classification Table
ISO Class 1: 0.2 um = 2 particles
ISO Class 2: 0.5 um =4 particles
ISOClass 3: 1.0 um = 8 particles
ISO Class 5: 5.0 um = 29 particles
Since it is possible to classify a cleanroom at one
particle size only, these low table values are
removed and cannot be used for classification.
95% Upper Confidence Limit
>1 & <10 sample points, 95% UCL should be 95% UCL removed completely

20. Thank-you..!
for your attention.
Have a wonderful dayahead…