EN 13610:2002

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Chemische Desinfektionsmittel - Quantitativer Suspensionsversuch zur Bestimmung der viruziden Wirkung gegenüber Bakteriophagen von chemischen Desinfektionsmitteln in den Bereichen Lebensmittel, und Industrie - Prüfverfahren und Anforderung (Phase 2, Stufe 1)Désinfectants chimiques - Essai quantitatif de suspension pour l'évaluation de l'activité virucide, vis a vis des bactériophages, des désinfectants chimiques utilisés dans les domaines de l'agro-alimentaire et de l'industrie - Méthode d'essai et prescriptions (phase 2, étape 1)Chemical disinfectants - Quantitative suspension test for the evaluation of virucidal activity against bacteriophages of chemical disinfectants used in food and industrial areas - Test method and requirements (phase 2, step 1)71.100.35Kemikalije za dezinfekcijo v industriji in domaChemicals for industrial and domestic disinfection purposesICS:Ta slovenski standard je istoveten z:EN 13610:2002SIST EN 13610:2003en01-november-2003SIST EN 13610:2003SLOVENSKI
STANDARD
EUROPEAN STANDARDNORME EUROPÉENNEEUROPÄISCHE NORMEN 13610December 2002ICS 11.080.20; 67.050; 71.100.35English versionChemical disinfectants - Quantitative suspension test for theevaluation of virucidal activity against bacteriophages ofchemical disinfectants used in food and industrial areas - Testmethod and requirements (phase 2, step 1)Désinfectants chimiques - Essai quantitatif de suspensionpour l'évaluation de l'activité virucide contre lesbactériophages des désinfectants chimiques utilisés dansle domaine de l'agro-alimentaire et dans l'industrie -Méthode d'essai et exigences (phase 2, étape 1)Chemische Desinfektionsmittel - QuantitativerSuspensionsversuch zur Bestimmung der viruzidenWirkung gegenüber Bakteriophagen von chemischenDesinfektionsmitteln in den Bereichen Lebensmittel, undIndustrie - Prüfverfahren und Anforderung (Phase 2, Stufe1)This European Standard was approved by CEN on 23 October 2002.CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this EuropeanStandard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such nationalstandards may be obtained on application to the Management Centre or to any CEN member.This European Standard exists in three official versions (English, French, German). A version in any other language made by translationunder the responsibility of a CEN member into its own language and notified to the Management Centre has the same status as the officialversions.CEN members are the national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece,Iceland, Ireland, Italy, Luxembourg, Malta, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and United Kingdom.EUROPEAN COMMITTEE FOR STANDARDIZATIONCOMITÉ EUROPÉEN DE NORMALISATIONEUROPÄISCHES KOMITEE FÜR NORMUNGManagement Centre: rue de Stassart, 36
B-1050 Brussels© 2002 CENAll rights of exploitation in any form and by any means reservedworldwide for CEN national Members.Ref. No. EN 13610:2002 ESIST EN 13610:2003

Test for validation of dilution-neutralization and molecular sieving methods.25Annex B (informative)
Neutralizers.33Annex C (informative)
Example of a typical test report.35Annex D (informative)
Information on the application and interpretation of European standards onchemical disinfectants and antiseptics.39Annex E (informative)
Example of plaques from lysates of phages P001 and P008.41Bibliography.44SIST EN 13610:2003

Lactococcuslactis subsp. lactis bacteriophage P001 and Lactococcus lactis subsp. lactis bacteriophage P008.Both phages shall be propagated on the host strain Lactococcus lactis subsp. lactis F7/2.The determined virucidal concentration of the tested product is suggested as being suitable for practical situationsof use.Where appropriate, additional specific virucidal activity against bacteriophages shall be determined under otherconditions of time, temperature, additional strains and interfering substances in accordance with (see 5.7.1) inorder to take into account intended specific use conditions.NOTEFor these additional conditions, the concentration defined as a result can be lower than the one obtained under theinitial test conditions of 20 °C, 15 min, 2 selected bacteriophage reference strains.5 Test methods5.1 Principle5.1.1A test suspension of bacteriophages in a solution of interfering substances is added to a prepared sampleof the product under test diluted in hard water.The mixture is maintained at 20 °C ± 1 °C for 15 min ± 10 s (required obligatory test conditions).At the end of the contact time, aliquots are taken and the virucidal activity against bacteriophages in this portion isimmediately neutralized or suppressed by a validated method. The method of choice is dilution-neutralization with avalidated neutralizer. If a suitable neutralizer is not available for a specific product, removal of the product bymolecular sieving (i.e. gel filtration) shall be used.The number of surviving bacteriophage particles and the number of bacteriophage particles in the test suspensionare determined from appropriate dilution series with a factor of 10 prepared in medium in test tubes.5.1.2Additional and optional exposure times, temperatures and interfering substances are specified (see 5.7.1).NOTE 1The test described is based on an assessment (under specific conditions) which gives a reduction of at least99,99 % (4 lg) of the infectivity of the different phages after different contact times.NOTE 2For principal reasons, the result of an inactivation applied to a viral population is not necessarily equal to 100 %: i.e.,one cannot conclude that there is a 100 % inactivation when on conducting the experiment no infectious phage are found withina limit number of sampling.5.2 Material and reagents5.2.1 Test organismsThe virucidal activity against bacteriophages shall be evaluated using the two following bacteriophage strains :SIST EN 13610:2003

1)DSM 4262, DSM 10567 and DSM 4366 are the collection numbers of bacteriophage and bacterial strains supplied by theDSMZ (Deutsche Sammlung von Mikroorganismen und ZellKulturen). This information is given for the convenience of users ofthis standard and does not constitute an endorsement by CEN of the culture collection named. Corresponding strains suppliedby other culture collections may be used if they can be shown to lead to the same results.SIST EN 13610:2003

2)Analytical quality of cross-linked dextran beads for molecular sieving (i.e. gel filtration). Sephadex® G-25 is an example of asuitable product available commercially. This information is given for the convenience of users of this standard and does notconstitute an endorsement by CEN of this product.SIST EN 13610:2003

1,0 %.5.2.2.14.3 Skim milkPrepare reconstituted skim milk (1,5 % fat content) for the test conditions as follows:¾ reconstitute skim milk powder, guaranteed free of antibiotics or additives, at a rate of 100 g/l of water (see5.2.2.2);¾ sterilize by steaming at 100 °C on 3 successive days (30 min each) and leave between steamings at roomtemperature.Do not leave between subsequent steamings in the refrigerator !NOTEUndiluted skim milk is used for maintenance of the bacterial host strain (see 5.4.1).Alternatively, sterilize at (3 0
115+) °C for 15 min.To obtain a volume fraction of 10 % working solution, dilute 1 part of skim milk with 9 parts of sterile water(see 5.2.2.2) which is required as an optional interfering substance for the phage suspension test (see 5.7.2).Store the volume fraction of 10 % skim milk at 4 °C to 8 °C.The final concentration of the skim milk in the test procedure (see 5.7.1) shall be a volume fraction of 1 %.5.3 Apparatus and glassware5.3.1 GeneralSterilize all glassware and parts of apparatus that will come into contact with the culture media and reagents or thesample, except those which are supplied sterile, by one of the following methods:a) in the autoclave (see 5.3.2.1) by maintaining it at (3 0
121+) °C for a minimum holding time of 15 min;b) in the dry heat sterilizer (see 5.3.2.1) by maintaining it at 180 °C for a minimum holding time of 30 min, at170 °C for a minimum holding time of 1 h or at 160 °C for a minimum holding time of 2 h.5.3.2Usual microbiological laboratory equipment 3) - in particular the following:5.3.2.1Apparatus for sterilization:a) for moist heat sterilization, an autoclave capable of being maintained at (3 0
121+) °C for a minimum holdingtime of 15 min;b) for dry heat sterilization, a hot air oven capable of being maintained at 180 °C for a minimum holding time of30 min, at 170 °C for a minimum holding time of 1 h or at 160 °C for a minimum holding time of 2 h.5.3.2.2Water baths capable of being controlled at 20 °C ± 1 °C, at additional test temperatures ± 1 °C (see5.7.1), and at 4 °C ± 1 °C for cooling down melted M17 top agar (see 5.5.2 and 5.5.3).5.3.2.3Incubator, capable of being controlled at 30 °C ± 1 °C.
3)Disposable equipment is an acceptable alternative to reusable glassware.SIST EN 13610:2003

4)Vortexâ is an example of a suitable product available commercially. This information is given for the convenience of usersof this standard and does not constitute an endorsement by CEN of this product.SIST EN 13610:2003

5)Confluent lysis usually occurs on agar plates derived from the 10-3 or 10-4 dilution step.6)PFU/ml = Plaque-forming units per ml.SIST EN 13610:2003

7)Analytical quality, non-hydrolized in accordance with European Pharmacopoeia volume 1. TWEEN 80® is an example of asuitable product available commercially. This information is given for the convenience of users of this standard and does notconstitute an endorsement by CEN of this product.SIST EN 13610:2003
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