ASTM E1838-17 Standard Test Method for Determining the Virus-Eliminating Effectiveness of Hygienic Handwash and Handrub Agents Using the Fingerpads of Adults.pdf

1、Designation: E1838 17Standard Test Method forDetermining the Virus-Eliminating Effectiveness of HygienicHandwash and Handrub Agents Using the Fingerpads ofAdults1This standard is issued under the fixed designation E1838; the number immediately following the designation indicates the year oforiginal

2、adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.INTRODUCTIONHands play an important role in the spread of many viruses. Thus, pr

3、oper and regular hand hygieneis crucial in preventing such spread, particularly in health-care settings, day-care centers, andfood-handling establishments. Many viruses that are known to spread through contaminated hands canremain infectious for several hours on human hands, and also may be more res

4、istant than the bacteriacommonly used to evaluate the microbicidal activity of handwash and handrub agents (1, 2, 3, 4).2Contaminated hands also can readily transfer infectious virus to other surfaces (1, 2, 3). Handantisepsis has been shown to interrupt the spread of viral infections (5, 6, 7, 8, 9

5、). This test methodis to assess the virus-eliminating potential of handwash and handrub agents in vivo.1. Scope1.1 Human skin is not known to carry viruses as a part of itsresident microbiota, with the notable exception of papillomaviruses (10). Hands transiently contaminated with viruses can,howeve

6、r, act as vehicles for the spread of many types of viralinfections. Hand hygiene is meant to reduce the load of virusesand other transient microorganisms on hands, thereby reducingthe risk of disease transmission. Such reductions in the virusload may be due to a combination of virus inactivation and

7、mechanical removal of infectious virus from the skin.1.2 This test method is designed to determine the compara-tive virus-eliminating effectiveness of microbicidal or non-microbicidal formulations. This test method is not meant foruse with surgical hand scrubs or preoperative skin preps.NOTE 1The te

8、st method should be performed by persons with trainingin virology in facilities designed and equipped for work with infectiousagents at biosafety level 2 (11).1.3 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.4 This standard

9、 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 limitations prior to use.1.5 This international standard

10、was developed in accor-dance with internationally recognized principles on standard-ization established in the Decision on Principles for theDevelopment of International Standards, Guides and Recom-mendations issued by the World Trade Organization TechnicalBarriers to Trade (TBT) Committee.2. Refere

11、nced Documents2.1 ASTM Standards:3E2011 Test Method for Evaluation of Hygienic Handwashand Handrub Formulations for Virus-Eliminating ActivityUsing the Entire HandE2276 Test Method for Determining the Bacteria-Eliminating Effectiveness of Hygienic Handwash andHandrub Agents Using the Fingerpads of A

12、dultsE2613 Test Method for Determining Fungus-EliminatingEffectiveness of Hygienic Handwash and HandrubAgentsUsing Fingerpads of Adults3. Terminology3.1 Definitions of Terms Specific to This Standard:3.1.1 Health-care personnel (HCP), npersons who aredirectly related to provision of health care serv

13、ices. It includes1This test method is under the jurisdiction of ASTM Committee E35 onPesticides, Antimicrobials, and Alternative Control Agents and is the directresponsibility of Subcommittee E35.15 on Antimicrobial Agents.Current edition approved April 1, 2017. Published June 2017. Originallyapprov

14、ed in 1996. Last previous edition approved in 2010 as E1838 10. DOI:10.1520/E1838-17.2The boldface numbers in parentheses refer to the list of references at the end ofthis standard.3For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.or

15、g. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United StatesThis international standard was developed in accordance with intern

16、ationally recognized principles on standardization established in the Decision on Principles for theDevelopment of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.1all paid and unpaid persons working in health-car

17、e settings,such as physicians, nurses, nursing assistants, therapists,technicians, emergency medical service personnel, dentalpersonnel, pharmacists, laboratory personnel, autopsypersonnel, students, trainees, and contractual staff, etc., whohave the potential to get themselves exposed to patients a

18、ndinfectious materials.3.1.2 hygienic (health-care personnel) handwash agents,nagents generally used for handwashing by personnel inhospitals, other health-care facilities, day-care centers, nursinghomes, and food-handling establishments should be safe forrepeated use, nonirritating, fast-acting, an

19、d efficient in elimi-nating transient microorganisms from intact skin.3.1.3 nonmedicated soap, na soap or detergent that is mildto the skin and does not contain any microbicidal chemicals.3.1.4 soil(organic) load, na solution of one or moreorganic and/or inorganic substances added to the suspension

20、ofthe test organism to simulate the presence of body secretions,excretions or other extraneous substances.3.1.5 virus-eliminating (killing/removing) agent, nanyagent that rids hands of viruses by either killing them on theskin or by dislodging them for subsequent wash-off.3.1.6 virus inactivating ag

21、ent, nany agent that renders avirus noninfectious.4. Summary of Test Method4.1 This test method is conducted on a group of adultsubjects who have provided informed consent and the skin ofwhose hands has been determined to be free from any apparentdamage. The subjects are to refrain from using any pr

22、oductscontaining antimicrobial agents for at least one week prior tothe test.Aknown volume of the test virus suspension is placedon a demarcated area on each fingerpad and the inoculumallowed to dry. The contaminated area then is exposed to testor control agent or a vehicle (for example, standard ha

23、rdwater), and rubbed with a randomly chosen fingerpad from theopposite hand for the desired contact time. Virus remaining onthe fingerpads is then eluted and the eluates titrated forinfectious virus along with the required controls. The infec-tious units from the two thumbpads or the pair of the fin

24、gerpadsthat were involved in a single treatment will be averaged.Percent or log10reductions, or both, in the levels of infectiousvirus after treatment with the test or control agents are thendetermined. If two different formulations are being comparedin the same test, one of them may be designated a

25、s a reference.If desired, one also may use tap water in parallel with the hardwater control to determine the influence of water hardness onthe test products virus-eliminating activity.5. Significance and Use5.1 This in vivo procedure is designed to test the ability ofhygienic handwash and handrub ag

26、ents to reduce levels ofselected infectious viruses from experimentally contaminatedfingerpads of adults. Since the two thumbpads and all eightfingerpads can be contaminated with virus and used in a giventest, it allows for the incorporation of a wet inoculum inputcontrol, dried virus recovery contr

27、ol, and up to three replicatesto assess the virus-eliminating efficiency of a test or controlagent, or a vehicle material. No more than 100 L of the virussuspension are required to complete one test.5.2 This test method is designed to be performed by atrained individual, who is responsible for choos

28、ing the appro-priate host system for the test virus and applying the techniquesnecessary for propagation and maintenance of host and testvirus. For a reference text, refer to Lennette et al (12).5.3 Whereas the method described here relates to testingwith viruses of human origin, it can be readily a

29、dapted to workwith animal pathogenic viruses as well as bacteriophages.Standard methods for working with bacteria (Test MethodE2276) and fungi (Test Method E2613) are also available.5.4 Infectious microorganisms left on hands after washingcan be reduced further by drying the washed hands with paper,

30、cloth, or warm air (13).Astep for the drying of fingerpads afterexposure to the control or test product, therefore, has not beenincluded to avoid virus removal by the drying process itself.5.5 This test method is not meant for use with surgical handscrubs or preoperative skin preps.5.6 The level of

31、viable virus in the dried inocula the controlfingerpads should not be less than 104infectious units whichwould permit the detection of up toa4log10reduction in theinfectivity titer of the virus by the test product under theconditions of this test method.6. Equipment and Apparatus6.1 Laminar Flow Cab

32、inetA Class II biological safetycabinet is required for virus work. The procedures for theproper maintenance and use of such cabinets are given in Ref(11).6.2 IncubatorAn incubator at 36 6 1C is needed forgrowing host cells and for incubating virus-infected cultures. Ifan open system is used for cel

33、l culture, a CO2incubator will berequired.6.3 Positive Displacement PipetteA pipette and pipettetips that accurately can dispense 10-L volumes.6.4 SterilizerAny steam sterilizer suitable for processingcell culture media and reagents is acceptable. The steamsupplied to the sterilizer must be free fro

34、m additives toxic tocell cultures.6.5 Filter Sterilization SystemA membrane or cartridgefiltration system (0.22-m pore diameter) is required forsterilizing heat-sensitive media and solutions.6.6 FreezersA freezer at 206 2C is required for thestorage of fetal bovine serum and other additives for cell

35、culture media. A second freezer at 70C or lower is requiredto store viruses6.7 RefrigeratorA refrigerator at 46 2C for storage ofprepared cell culture media and reagents.6.8 TimerAny stopwatch that can be read in minutes andseconds.6.9 Handwashing SinkA sink of sufficient size to permitsubjects to w

36、ash hands without touching hands to sink surface.E1838 1726.9.1 Water Faucet(s), to be located above the sink at aheight that permits the hands to be held higher than the elbowduring the washing procedure. Faucets with electronic sensorsor those that are wrist-, elbow-, knee-, or foot-operated arepr

37、eferred to avoid recontamination of the washed hands.6.9.2 Tap Water Temperature Regulator and TemperatureMonitor, to monitor and regulate water temperature at 40 62C.6.10 Liquid Nitrogen Storage for CellsA proper liquidnitrogen container and liquid nitrogen for cryopreservation ofthe stocks of cell

38、 lines.6.11 Inverted MicroscopeAn inverted microscope with10 eye pieces and 5, 10, and 40 objectives.7. Materials and Reagents7.1 Serological PipettesSterile reusable or single-use pi-pettes of 10.0, 5.0, and 1.0-mL capacity.7.2 Cell Culture FlasksPlastic flasks of 25 or 75-cm2capacity for culturing

39、 cells and for preparing virus pools.NOTE 2Each flask for growing cell monolayers can be reused ten ormore times before being discarded.NOTE 3Plastic cell culture ware may be purchased from mostlaboratory supply houses.7.3 Cell Culture Plates, 6-well2.0 mL per well eluent (see7.8) shall be added. Th

40、is is used for virus elution from eachthumbpad and fingerpad.NOTE 4Alternatively, small dishes (for example, 35mm diameter)may be used for virus elution.7.4 Cell Culture Media and SupplementsCulture mediaand the types and ratios of supplements will vary depending onthe cell line. Eagles minimal esse

41、ntial medium (EMEM) with5 to 10 % fetal bovine serum (virus- and mycoplasma-tested) isused for growing a wide variety of cells (see Note 5).NOTE 5Materials and reagents for cell culture may be purchased frombiological supply houses.7.5 Soil Load:7.5.1 Bovine Serum, at a final concentration of 5 % in

42、 thevirus inoculum (see Note 6).7.5.2 A Yeast extract/BSA/Mucin tripartite soil load, as analternative to serum. Add 0.5 g of yeast extract to 10 mL ofphosphate buffer.Add 0.5 g of bovine serum albumin (BSA) to10 mL of phosphate buffer. Add 0.04 g of bovine mucin to 10mL of phosphate buffer. Prepare

43、 the stock solutions separatelyand sterilize by passage through a 0.22 m pore diametermembrane filter, aliquot and store at either 462C or2062C. To obtain a 500-L inoculum of the test inoculum,add to 340 L of the microbial suspension 25 L BSA, 100 Lmucin and 35 L of yeast extract stock solutions. Th

44、is mixturecontains approximately2goftotal protein/L, which is approxi-mately equivalent to the protein content of a 5 % solution offetal bovine serum.NOTE 6Bovine serum is unsuitable for use as an organic load whenworking with rotaviruses because of its rotavirus inhibitory and trypsin-neutralizing

45、activity.7.6 Standard Hard WaterStandard hard water prepared inaccordance with AOAC 960.09 E and F (14) at a hardness of200 ppm as calcium carbonate is used for dilution of testsubstance, as the control solution to determine the baselinelevel of virus elimination, and to rinse the fingerpads afterex

46、posure to the test product. The standard hard water and tapwater (if used) must first be tested to ensure that they do nothave any virucidal activity against the test virus(es).NOTE 7The quality and disinfectant (for example, chlorine) residualin tap water can vary from site to site and at different

47、 times at the samesite. The use of standard hard water, therefore, is recommended here toavoid variations in results due to differences in tap water quality.7.7 Test productTwo separate manufacturers lots of thetest product may be tested. For handwash products that areused with water, prepare a 25 %

48、 solution by adding 1 partproduct to 3 parts standard hard water. This dilution isnecessary because water is used when the product is applied.7.8 Eluent for Virus Recovery from FingerpadsMinimumEssential Medium (MEM) + 2 % Fetal Bovine Serum (FBS),or Earles balanced salt solution (EBSS) with a pH of

49、 7.2 7.4,or equivalent.7.9 Diluent for Virus TitrationSame as the Eluent forVirus Recovery from Fingerpads.7.10 Plastic VialsSterile screw-capped 2.0-mL vials withan inside diameter of about 8 mm are required for demarcationof the fingerpads and to hold various test solutions.7.11 Miscellaneous Laboratory WareAutomatic pipettes,pipette tips, plastic vials for storing cell and virus stocks,dilution tubes, cluster plates, or flasks for virus titration.8. Test Viruses and Cell Cultures8.1 See Appendix X1 for rec