1、Designation:E148212Standard Practice forUse of Gel Filtration Columns for Cytotoxicity Reductionand Neutralization1This standard is issued under the fixed designation E1482;the number immediately following the designation indicates the year oforiginal adoption or,in the case of revision,the year of
2、last revision.A number in parentheses indicates the year of last reapproval.Asuperscript epsilon()indicates an editorial change since the last revision or reapproval.1.ScopeNOTE1The title was formerly Standard Test Method for Neutraliza-tion of Virucidal Agents in Virucidal Efficacy Evaluations.1.1
3、This practice is intended to be used to reduce thecytotoxic level of the virus-test product mixture prior toassaying for viral infectivity.It is used in conjunction withevaluations of the virucidal efficacy of disinfectant solutions,wipes,trigger sprays,or pressurized disinfectant spray prod-ucts in
4、tended for use on inanimate,nonporous environmentalsurfaces.This practice may also be used in the evaluation ofhygienic handwashes/handrubs,or for other special applica-tions.The practice may be employed with all viruses and hostsystems.1.2 This practice should be performed only by personstrained in
5、 virology techniques.1.3 This practice utilizes gel filtration technology.Theeffectiveness of the practice is dependent on the ratio of gel bedvolume to sample size and uniformity in the preparation ofcolumns as well as the conditions of entrifugation.Theeffectiveness of this practice is maximized b
6、y investigatorpractice and experience with gel filtration techniques.1.4 This practice will aid in the reduction,but not necessar-ily elimination,of test product toxicity while preserving thetiter of the input virus.1.5 The values stated in SI units are to be regarded asstandard.No other units of me
7、asurement are included in thisstandard.1.6 This standard does not purport to address all of thesafety concerns,if any,associated with its use.It is theresponsibility of the user of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limi
8、tations prior to use.2.Referenced Documents2.1 ASTM Standards:2E1052 Test Method to Assess the Activity of Microbicidesagainst Viruses in SuspensionE1053 Test Method to Assess Virucidal Activity of Chemi-cals Intended for Disinfection of Inanimate,NonporousEnvironmental Surfaces3.Summary of Test Met
9、hods3.1 After the exposure of a virus to a test product(orhandwash/rub product),the virus-product suspension is addedto a column of Sephadex3LH-60,Sephadex3LH-20,orSephacryl3S-1000 Superfine.The column(encased within asterile centrifuge tube in order to capture the filtrate)is placedin a centrifuge
10、and centrifuged to separate the virus from thetest product by gel filtration.Alternatively,samples may behand-plunged using a syringe plunger.The filtrate(the columnflow-through which contains the virus)is assayed in theappropriate host system.The untreated virus control suspen-sion is gel-column fi
11、ltered,using the same methods/techniques,and the virus titer of the filtrate is determined by assay ofinfectivity.The residual cytotoxicity of the disinfectant isdetermined by gel filtration of the test product control under thesame conditions as those which were used in the test.Resultsfor the viru
12、s inactivation and test product cytotoxicity ofgel-column filtrates are recorded in the same manner asdescribed in Test Methods E1052 and E1053.The gel-columnfiltration procedures described in this practice are a modifica-tion of the method of Blackwell and Chen.4NOTE2A limitation of utilizing colum
13、ns in virological assays is thatthey are unable to effectively neutralize all actives.Prior to testing,ensure1This practice is under the jurisdiction of ASTM Committee E35 on Pesticides,Antimicrobials,and Alternative Control Agentsand is the direct responsibility ofSubcommittee E35.15 on Antimicrobi
14、al Agents.Current edition approved Oct.1,2012.Published November 2012.Originallyapproved in 1992.Last previous edition approved in 2004 as E1482 04).DOI:10.1520/E1482-12.2For referenced ASTM standards,visit the ASTM website,www.astm.org,orcontact ASTM Customer Service at serviceastm.org.For Annual B
15、ook of ASTMStandards volume information,refer to the standards Document Summary page onthe ASTM website.3Sephadex is a registered trademark of Amersham Biosciences.The sole sourceof supply of the apparatus known to the committee at this time is AmershamBiosciences.If you are aware of alternative sup
16、pliers,please provide this informa-tion to ASTM International Headquarters.Your comments will receive carefulconsideration at a meeting of the responsible technical committee,1which you mayattend.4Blackwell,H.H.,and Chen,J.H.S.,“Effects of Various Germicidal Chemicalson H.EP.2 Cell Culture and Herpes simplex Virus,”Journal of the AOAC,Vol 53,1970,pp.12291236.Copyright ASTM International,100 Barr Harbor Drive,PO Box C700,West Conshohocken,PA 19428-2959.United States1 the effectiveness of gel-filt
