5.1.10. Guidelines for using the test for bacterial endotoxins EUROPEAN PHARMACOPOEIA 7.

meets the requirements of the European Pharmacopoeia. A 01/2010:50110

manufacturer is neither obliged to carry out such tests nor
precluded from using modifications of, or alternatives to, the
stated method, provided he is satisfied that, if tested by the 5.1.10. GUIDELINES FOR USING THE
official method, the material in question would comply with the TEST FOR BACTERIAL ENDOTOXINS
requirements of the European Pharmacopoeia.
1. INTRODUCTION
Endotoxins from gram-negative bacteria are the most common
cause of toxic reactions resulting from contamination of
PRECAUTIONS AGAINST MICROBIAL CONTAMINATION pharmaceutical products with pyrogens ; their pyrogenic activity
Aseptic conditions for performance of the test can be achieved is much higher than that of most other pyrogenic substances.
using, for example, a class A laminar-air-flow cabinet located These endotoxins are lipo-polysaccharides. Although there are
within a class B clean room, or an isolator a small number of pyrogens which possess a different structure,
the conclusion is generally justified that the absence of bacterial
endotoxins in a product implies the absence of pyrogenic
components, provided the presence of non-endotoxin pyrogenic
GUIDANCE TO MANUFACTURERS substances can be ruled out.
The level of assurance provided by a satisfactory result of a The presence of endotoxins in a product may be masked by
test for sterility (the absence of contaminated units in the factors interfering with the reaction between the endotoxins
sample) as applied to the quality of the batch is a function of and the amoebocyte lysate. Hence, the analyst who wishes to
the homogeneity of the batch, the conditions of manufacture replace the rabbit pyrogen test required in a pharmacopoeial
and the efficiency of the adopted sampling plan. Hence for monograph by a test for bacterial endotoxins has to demonstrate
the purpose of this text a batch is defined as a homogeneous that a valid test can be carried out on the product concerned ;
collection of sealed containers prepared in such a manner that this may entail a procedure for removing interfering factors.
the risk of contamination is the same for each of the units As indicated in the test for bacterial endotoxins (2.6.14),
contained therein. information must be available on the 2 following aspects before
a test on a sample can be regarded as valid.
In the case of terminally sterilised products, physical proofs,
— The suitability of the material to be used for the test has to be
biologically based and automatically documented, showing
established. The absence of endotoxins in the water for BET
correct treatment throughout the batch during sterilisation are
and in the other reagents must be assured and the sensitivity
of greater assurance than the sterility test. The circumstances
of the amoebocyte lysate must be checked to confirm the
in which parametric release may be considered appropriate
sensitivity declared by the manufacturer.
are described under 5.1.1. Methods of preparation of sterile
products. The method of media-fill runs may be used to — As the product to be examined may interfere with the test,
evaluate the process of aseptic production. Apart from that, the the sensitivity of the amoebocyte lysate is determined in
sterility test is the only analytical method available for products the presence and in the absence of the product under
prepared under aseptic conditions and furthermore it is, in all examination. There must be no significant difference
cases, the only analytical method available to the authorities between the 2 sensitivity values.
who have to examine a specimen of a product for sterility. The text 2.6.14. Bacterial endotoxins indicates methods for
removing interfering factors ; in the case of interference, another
The probability of detecting micro-organisms by the test for test must be carried out after such a method has been applied
sterility increases with their number present in the sample to check whether the interference has indeed been neutralised
tested and varies according to the readiness of growth of or removed.
micro-organism present. The probability of detecting very low This general chapter explains the reasons for the requirements
levels of contamination even when it is homogenous throughout in the test for bacterial endotoxins, then deals with the reading
the batch is very low. The interpretation of the results of the test and interpretation of the results.
for sterility rests on the assumption that the contents of every
container in the batch, had they been tested, would have given Substitution of the rabbit pyrogen test required in a
the same result. Since it is manifest that every container cannot pharmacopoeial monograph by an amoebocyte lysate test
be tested, an appropriate sampling plan should be adopted. In constitutes the use of an alternative method of analysis and
the case of aseptic production, it is recommended to include hence requires validation ; some guidance on how to proceed is
samples filled at the beginning and at the end of the batch and given in section 11.
after significant intervention. The reference method for bacterial endotoxins is stated in the
monograph on a given product ; where no method is stated,
method A is the reference method. If a method other than the
reference method is to be used, the analyst must demonstrate
OBSERVATION AND INTERPRETATION OF RESULTS that the method is appropriate for this product and gives a
Conventional microbiological/biochemical techniques are result consistent with that obtained with the reference method
generally satisfactory for identification of micro-organisms (see also Section 13).
recovered from a sterility test. However, if a manufacturer
wishes to use condition (d) as the sole criterion for invalidating 2. METHOD
a sterility test, it may be necessary to employ sensitive typing The addition of endotoxins to amoebocyte lysate may result in
techniques to demonstrate that a micro-organism isolated turbidity, precipitation or gelation (gel-clot) ; only the gel-clot
from the product test is identical to a micro-organism isolated method was used in the Pharmacopoeia as an evaluation
from the test materials and/or the testing environment. While criterion in the first type of test for bacterial endotoxins. The
routine microbiological/biochemical identification techniques advantage was the simplicity of basing the decision to pass or
can demonstrate that 2 isolates are not identical, these fail the product under examination on the absence or presence
methods may not be sufficiently sensitive or reliable enough of a gel-clot, visible with the naked eye. The quantitative
to provide unequivocal evidence that 2 isolates are from the methods described as methods C, D, E and F were developed
same source. More sensitive tests, for example molecular typing later : they require more instrumentation, but they are easier to
with RNA/DNA homology, may be necessary to determine that automate for the regular testing of large numbers of samples of
micro-organisms are clonally related and have a common origin. the same product.

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EUROPEAN PHARMACOPOEIA 7.0 5.1.10. Guidelines for using the test for bacterial endotoxins

Endotoxins may be adsorbed onto the surface of tubes endotoxin limit and that a positive result means that the lysate
or pipettes made from certain plastics or types of glass. detected an endotoxin concentration equal to or greater than
Interference may appear due to the release of substances from the endotoxin limit? This dilution depends on the endotoxin
plastic materials. Hence, the materials used should be checked ; limit and on the sensitivity of the lysate : it is called the Maximum
subsequent batches of tubes or pipettes may have a slightly Valid Dilution (MVD) and its value may be calculated using the
different composition, and therefore the analyst is advised to following expression :
repeat such tests on starting with new batches of materials.
The decision to use the test for bacterial endotoxins as a limit
test implies first that a threshold endotoxin concentration
must be defined for the product to be tested, and second that Concentration of test solution :
the objective of the test is to know whether the endotoxin
concentration in the product under examination is below or — mg/mL if the endotoxin limit is specified by mass (IU/mg),
above this threshold. The quantitative methods C, D, E and F — Units/mL if the endotoxin limit is specified by unit of
make it possible to determine the endotoxin concentration in biological activity (IU/Unit),
the sample under examination, but for compliance with the
— ml/mL if the endotoxin limit is specified by volume (IU/mL).
Pharmacopoeia and in routine quality control the final question
is whether or not this concentration exceeds a defined limit. λ = the labelled lysate sensitivity in the gel-clot technique
In setting a threshold concentration of endotoxin for the (IU/mL) or the lowest concentration used in the
product to be tested, due attention should be paid to the dose standard curve of the turbidimetric or chromogenic
of the product : the threshold should be set so as to ensure that techniques.
as long as the endotoxin concentration in the product remains When the value of the maximum valid dilution is not a whole
below this threshold even the maximal dose administered by the number, a convenient whole number smaller than the MVD
intended route per hour does not contain sufficient endotoxin may be used for routine purposes (which means preparing a
to cause a toxic reaction. solution of the product which is less diluted than the MVD
When the endotoxin concentration in the product exactly equals indicates). In this case, a negative result indicates that the
the threshold value, gelation will occur, as is the case when endotoxin concentration of the product lies below the limit
the endotoxin concentration is much higher, and the product value. However, when the endotoxin concentration of the
will fail the test, because the all-or-none character of the test product in such a test is less than the endotoxin limit but high
makes it impossible to differentiate between a concentration enough to make the reaction with the lysate result in a clot, the
exactly equal to the threshold concentration and one that is test may be positive under these conditions. Hence, when a test
higher. It is only when no gelation occurs that the analyst with this ‘convenient’ dilution factor is positive, the product
may conclude that the endotoxin concentration is below the should be diluted to the MVD and the test should be repeated.
threshold concentration. In any case of doubt or dispute the MVD must be used.
For products in the solid state, this threshold concentration This stresses the importance of the confirmation of the
of endotoxin per mass unit or per International Unit (IU) of sensitivity of the lysate.
product has to be translated into a concentration of endotoxin
per millilitre of solution to be tested, as the test can only be Example
carried out on a solution. The case of products that already exist A 50 mg/mL solution of phenytoin sodium (intended for
in the liquid state (such as infusion fluids) is discussed below. intravenous injection) has to be tested. Determine the MVD,
Endotoxin limit : the endotoxin limit for active substances given the following variables :
administered parenterally, defined on the basis of dose, is equal M = maximum human dose = 15 mg per kilogram of
to : body mass,
c = 50 mg/mL,
K = 5 IU of endotoxin per kilogram of body mass,
K = threshold pyrogenic dose of endotoxin per kilogram λ = 0.4 IU of endotoxin per millilitre.
of body mass,
M = maximum recommended bolus dose of product per
kilogram of body mass.
When the product is to be injected at frequent intervals or For routine tests on this product, it may be expedient to dilute
infused continuously, M is the maximum total dose administered 1 mL of the solution to be tested to 20 mL (MVD/2 rounded to
in a single hour period. the next lower whole number). However, if this test result is
The endotoxin limit depends on the product and its route of positive the analyst will have to dilute 1 mL to 41.67 mL and
administration and is stated in the monograph. Values for K are repeat the test. A dilution to 41.67 mL is also necessary when
suggested in Table 5.1.10.-1. the test is performed to settle a dispute.
For other routes, the acceptance criterion for bacterial 3. REFERENCE MATERIAL
endotoxins is generally determined on the basis of results
obtained during the development of the preparation. Endotoxin standard BRP is intended for use as the reference
preparation. It has been assayed against the WHO International
Table 5.1.10.-1 Standard for Endotoxin and its potency is expressed in
Route of administration K (IU of endotoxin per kilogram of body mass) International Units of endotoxin per ampoule. The International
Unit of endotoxin is defined as the specific activity of a defined
Intravenous 5.0
mass of the International Standard.
Intravenous, for 2.5 For routine purposes, another preparation of endotoxin may be
radiopharmaceuticals
used, provided it has been assayed against the International
Intrathecal 0.2
Standard for Endotoxin or the BRP and its potency is expressed
Which dilution of the product is to be used in the test to in International Units of endotoxin.
obtain maximal assurance that a negative result means that NOTE : 1 International Unit (IU) of endotoxin is equal to
the endotoxin concentration of the product is less than the 1 Endotoxin Unit (E.U.).

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4. WATER FOR BET Methods C and D. If the nature of the product to be analysed
Testing the absence of endotoxin in this reagent by a technique shows interference which cannot be removed by classical
derived from the rabbit pyrogen test was rejected for practical methods, it may be possible to determine the standard curve in
and theoretical reasons : the same type of product freed from endotoxins by appropriate
treatment or by dilution of the product. The endotoxins test is
— the rabbit test is not sensitive enough to detect endotoxin in then carried out by comparison with this standard curve.
water for BET intended for tests on products with a very low
endotoxin limit; Ultrafiltration with cellulose triacetate asymmetric membrane
filters has been found to be suitable in most cases. The
— the relatively low precision of the rising temperature response filters should be properly validated, because under some
in rabbits would call for many replications in rabbits ; circumstances cellulose derivatives (β-D-glucans) can cause false
— the terms ‘pyrogens’ and ‘endotoxins’ denote groups of positive results.
entities that do not coincide completely. Polysulfone filters have been found to be unsuitable because
The text 2.6.14. Bacterial endotoxins indicates that methods false positive results had been obtained by some users.
other than triple distillation may be used to prepare water
for BET. Reverse osmosis has been used with good results ; 9. THE PURPOSE OF THE CONTROLS
some analysts may prefer to distil the water more than 3 times. The purpose of the control made up with water for BET and the
Whatever method is used, the resultant product must be free of reference preparation of endotoxin at twice the concentration of
detectable endotoxins. the labelled lysate sensitivity is to verify the activity of the lysate
at the time and under the conditions of the test. The purpose
5. pH OF THE MIXTURE of the negative control is to verify the absence of a detectable
In the test for bacterial endotoxins, optimum gel-clot occurs for concentration of endotoxin in water for BET.
a mixture at pH 6.0-8.0. However, the addition of the lysate to The positive control, which contains the product to be examined
the sample may result in a lowering of the pH. at the concentration used in the test, is intended to show
the absence of inhibiting factors at the time and under the
6. VALIDATION OF THE LYSATE conditions of the test.
It is important to follow the manufacturer’s instructions for the 10. READING AND INTERPRETATION OF THE RESULTS
preparation of the solutions of the lysate.
Minute amounts of endotoxin in the water for BET, or in any
The positive end-point dilution factors in gel-clot methods A other reagent or material to which the lysate is exposed during
and B are converted to logarithms. The reason is that if the the test, may escape detection as long as they do not reach
frequency distribution of these logarithmic values is plotted, the sensitivity limit of the lysate. However, they may raise the
it usually approaches a normal distribution curve much more amount of endotoxin in the solution containing the product
closely than the frequency distribution of the dilution factors under examination to just above the sensitivity limit and cause
themselves ; in fact it is so similar that it is acceptable to use the
a positive reaction.
normal frequency distribution as a mathematical model and to
The risk of this happening may be reduced by testing the water
calculate confidence limits with Student’s t-test.
for BET and the other reagents and materials with the most
7. PRELIMINARY TEST FOR INTERFERING FACTORS sensitive lysate available, or at least one that is more sensitive
than the one used in the test on the product. Even then,
Some products cannot be tested directly for the presence of the risk of such a ‘false positive result’ cannot be ruled out
endotoxins because they are not miscible with the reagents, completely. It should be realised, however, that in this respect
they cannot be adjusted to pH 6.0-8.0 or they inhibit or activate the test design is ‘fail-safe’ in contrast to a test design permitting
gel formation. Therefore a preliminary test is required to check a false negative result, which could lead to the release of an
for the presence of interfering factors ; when these are found the unsatisfactory product, thus endangering the patient’s health.
analyst must demonstrate that the procedure to remove them
has been effective. 11. REPLACEMENT OF THE RABBIT PYROGEN TEST BY A
The object of the preliminary test is to test the null hypothesis TEST FOR BACTERIAL ENDOTOXINS
that the sensitivity of the lysate in the presence of the product Monographs on pharmaceutical products intended for
under examination does not differ significantly from the parenteral administration that may contain toxic amounts
sensitivity of the lysate in the absence of the product. A simple of bacterial endotoxins require either a test for bacterial
criterion is used in methods A and B : the null hypothesis is endotoxins or a rabbit pyrogen test. As a general policy :
accepted when the sensitivity of the lysate in the presence of — in any individual monograph, when a test is required, only
the product is at least 0.5 times and not more than twice the one test is included, either that for pyrogens or that for
sensitivity of the lysate by itself. bacterial endotoxins ;
A classical approach would have been to calculate the means of — in the absence of evidence to the contrary, the test for
the log dilution factor for the lysate sensitivity with and without bacterial endotoxins is preferred over the test for pyrogens,
the product and to test the difference between the 2 means with since it is usually considered to provide equal or better
Student’s t-test. protection to the patient;
The test for interfering factors in gel-clot methods A and B — before including a test for bacterial endotoxins in a
requires the use of a sample of the product in which no monograph, evidence is required that one of the tests
endotoxins are detectable. This presents a theoretical problem described in chapter 2.6.14 can be applied satisfactorily to
when an entirely new product has to be tested. Hence, a different the product in question ;
approach was designed for quantitative methods C, D, E and F. — the necessary information is sought from manufacturers ;
companies are invited to provide any validation data that
8. REMOVAL OF INTERFERING FACTORS they have concerning the applicability of the test for bacterial
The procedures to remove interfering factors must not increase endotoxins to the substances and formulations of interest ;
or decrease (for example, by adsorption) the amount of such data includes details of sample preparation and of any
endotoxin in the product under examination. The correct way procedures necessary to eliminate interfering factors ; in
of checking this is to apply the procedures to a spiked sample addition, any available parallel data for rabbit pyrogen testing
of the product, that is, a sample to which a known amount of that would contribute to an assurance that the replacement
endotoxin has been added, and then to measure the recovery of a rabbit pyrogen test by the test for bacterial endotoxin is
of the endotoxin. appropriate, must be provided.

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EUROPEAN PHARMACOPOEIA 7.0 5.1.10. Guidelines for using the test for bacterial endotoxins

Additional requirements are defined in the following sections. standards of the monographs would be achieved if the
official methods were used. In the event of doubt or dispute,
12. USE OF A DIFFERENT BACTERIAL ENDOTOXIN TEST the methods of analysis of the Pharmacopoeia are alone
FROM THAT PRESCRIBED IN THE MONOGRAPH authoritative.”
When a test for bacterial endotoxins is prescribed in a The following procedures are suggested for validating a method
monograph and none of the 6 methods (A to F) described in for bacterial endotoxins other than the one implied or indicated
chapter 2.6.14 is specified, then method A, the gel-clot method in the monograph.
limit test, has been validated for this product. If one of the other 13-1. The procedure and the materials and reagents used in the
methods (B to F) is specified, this is the one which has been method should be validated as described for the test concerned.
validated for this product.
13-2. The presence of interfering factors (and, if needed, the
13. VALIDATION OF ALTERNATIVE METHODS procedure for removing them) should be tested on samples
of at least 3 production batches. It should be borne in mind
Replacement of a rabbit pyrogen test by a bacterial endotoxin that methods D and E, using a chromogenic peptide, require
test, or replacement of a stated or implied method for bacterial reagents that are absent in methods A, B, C and F, and hence
endotoxins by another method, is to be regarded as the use of compliance of methods A, B, C or F with the requirements
an alternative method in the replacement of a pharmacopoeial for interfering factors cannot be extrapolated to method D or
test, as described in the General Notices : method E without further testing.
“The test and assays described are the official methods
upon which the standards of the Pharmacopoeia are based. 14. VALIDATION OF THE TEST FOR NEW PRODUCTS
With the agreement of the competent authority, alternative The procedures described under 13-1 and 13-2 should be applied
methods of analysis may be used for control purposes, to all new products intended for parenteral administration
provided that the methods used enable an unequivocal that have to be tested for the presence of bacterial endotoxins
decision to be made as to whether compliance with the according to the requirements of the Pharmacopoeia.

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 EUROPEAN PHARMACOPOEIA 7.0

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EUROPEAN PHARMACOPOEIA 7.0

5.2. GENERAL TEXTS ON

BIOLOGICAL PRODUCTS
5.2. General texts on biological products................................... 527 5.2.6. Evaluation of safety of veterinary vaccines and
5.2.1. Terminology used in monographs on biological immunosera .. ................................................................................. 536
products.. ......................................................................................... 527 5.2.7. Evaluation of efficacy of veterinary vaccines and
5.2.2. Chicken flocks free from specified pathogens for the immunosera.. .................................................................................. 538
production and quality control of vaccines.. ........................... 527 5.2.8. Minimising the risk of transmitting animal spongiform
5.2.3. Cell substrates for the production of vaccines for human encephalopathy agents via human and veterinary medicinal
use..................................................................................................... 530 products.. ......................................................................................... 539
5.2.4. Cell cultures for the production of veterinary 5.2.9. Evaluation of safety of each batch of veterinary vaccines
vaccines............................................................................................ 533 and immunosera.. .......................................................................... 547
5.2.5. Substances of animal origin for the production of
immunological veterinary medicinal products........................ 535

General Notices (1) apply to all monographs and other texts 525
 EUROPEAN PHARMACOPOEIA 7.0

526 See the information section on general monographs (cover pages)

EUROPEAN PHARMACOPOEIA 7.0 5.2.2. SPF chicken flocks for vaccines

5.2. GENERAL TEXTS ON Single harvest. Material derived on one or more occasions
from a single production cell culture inoculated with the same
BIOLOGICAL PRODUCTS working seed lot or a suspension derived from the working seed
lot, incubated, and harvested in a single production run.
Monovalent pooled harvest. Pooled material containing a
01/2008:50201 single strain or type of micro-organism or antigen and derived
corrected 6.0 from a number of eggs, cell culture containers etc. that are
processed at the same time.
5.2.1. TERMINOLOGY USED IN Final bulk vaccine. Material that has undergone all the steps
of production except for the final filling. It consists of one or
MONOGRAPHS ON BIOLOGICAL more monovalent pooled harvests, from cultures of one or more
PRODUCTS species or types of micro-organism, after clarification, dilution
or addition of any adjuvant or other auxiliary substance. It is
For some items, alternative terms commonly used in connection treated to ensure its homogeneity and is used for filling the
with veterinary vaccines are shown in parenthesis. containers of one or more final lots (batches).
Seed-lot system. A seed-lot system is a system according to Final lot (Batch). A collection of closed, final containers or
which successive batches of a product are derived from the other final dosage units that are expected to be homogeneous
same master seed lot. For routine production, a working seed and equivalent with respect to risk of contamination during
lot may be prepared from the master seed lot. The origin and filling or preparation of the final product. The dosage units
the passage history of the master seed lot and the working seed are filled, or otherwise prepared, from the same final bulk
lot are recorded. vaccine, freeze-dried together (if applicable) and closed in one
Master seed lot. A culture of a micro-organism distributed from continuous working session. They bear a distinctive number or
a single bulk into containers and processed together in a single code identifying the final lot (batch). Where a final bulk vaccine
operation in such a manner as to ensure uniformity and stability is filled and/or freeze-dried on several separate sessions, there
and to prevent contamination. A master seed lot in liquid form results a related set of final lots (batches) that are usually
is usually stored at or below − 70 °C. A freeze-dried master seed identified by the use of a common part in the distinctive number
lot is stored at a temperature known to ensure stability. or code ; these related final lots (batches) are sometimes referred
Working seed lot. A culture of a micro-organism derived from to as sub-batches, sub-lots or filling lots.
the master seed lot and intended for use in production. Working Combined vaccine. A multicomponent preparation formulated
seed lots are distributed into containers and stored as described so that different antigens are administered simultaneously.
above for master seed lots. The different antigenic components are intended to protect
Cell-bank system (Cell-seed system). A system whereby against different strains or types of the same organism and/or
successive final lots (batches) of a product are manufactured by different organisms. A combined vaccine may be supplied
culture in cells derived from the same master cell bank (master by the manufacturer either as a single liquid or freeze-dried
cell seed). A number of containers from the master cell bank preparation or as several constituents with directions for
(master cell seed) are used to prepare a working cell bank admixture before use.
(working cell seed). The cell-bank system (cell-seed system) is
validated for the highest passage level achieved during routine
production. 07/2010:50202
Master cell bank (Master cell seed). A culture of cells
distributed into containers in a single operation, processed 5.2.2. CHICKEN FLOCKS FREE FROM
together and stored in such a manner as to ensure uniformity
and stability and to prevent contamination. A master cell bank SPECIFIED PATHOGENS FOR THE
(master cell seed) is usually stored at − 70 °C or lower. PRODUCTION AND QUALITY CONTROL
Working cell bank (Working cell seed). A culture of cells OF VACCINES
derived from the master cell bank (master cell seed) and
intended for use in the preparation of production cell cultures. Where specified, chickens, embryos or cell cultures used for
The working cell bank (working cell seed) is distributed into the production or quality control of vaccines are derived from
containers, processed and stored as described for the master eggs produced by chicken flocks free from specified pathogens
cell bank (master cell seed). (SPF). The SPF status of a flock is ensured by means of the
system described below. The list of micro-organisms given is
Primary cell cultures. Cultures of cells obtained by trypsination based on current knowledge and will be updated as necessary.
of a suitable tissue or organ. The cells are essentially identical
to those of the tissue of origin and are no more than 5 in vitro A flock is defined as a group of birds sharing a common
passages from the initial preparation from the animal tissue. environment and having their own caretakers who have no
contact with non-SPF flocks. Once a flock is defined, no
Cell lines. Cultures of cells that have a high capacity for non-SPF birds are added to it.
multiplication in vitro. In diploid cell lines, the cells have
Each flock is housed so as to minimise the risk of contamination.
essentially the same characteristics as those of the tissue of
The facility in which the flock is housed must not be sited near
origin. In continuous cell lines, the cells are able to multiply
to any non-SPF flocks of birds with the exception of flocks
indefinitely in culture and may be obtained from healthy or
that are in the process of being established as SPF flocks and
tumoral tissue. Some continuous cell lines have oncogenic
that are housed in facilities and conditions appropriate to SPF
potential under certain conditions.
flocks. The SPF flock is housed within an isolator or in a
Production cell culture. A culture of cells intended for use in building with filtered air under positive pressure. Appropriate
production ; it may be derived from one or more containers of measures are taken to prevent entry of rodents, wild birds,
the working cell bank (working cell seed) or it may be a primary insects and unauthorised personnel.
cell culture. Personnel authorised to enter the facility must have no contact
Control cells. A quantity of cells set aside, at the time of virus with other birds or with agents potentially capable of infecting
inoculation, as uninfected cell cultures. The uninfected cells the flock. It is advisable for personnel to shower and change
are incubated under similar conditions to those used for the clothing or to wear protective clothing before entering the
production cell cultures. controlled facility.

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