Test No. 474: Mammalian Erythrocyte Micronucleus Test [electronic resource]

10 Pages·1997·0.28 MB·English

by  Organisation for Economic Co-operation and Development

474
Adopted:
21stJuly1997
OECDGUIDELINEFORTHETESTINGOFCHEMICALS
MammalianErythrocyteMicronucleusTest
INTRODUCTION
t1h.etestsTuhbestamnacmematolitahneicnhrvoivmoosmoicmreosnuocrletuhsetmeisttoitsicusaepdpafroartuthseodfeteecrtyitohnroobfladsatmsabgyeiannadluycseidsboyf
erythrocytesassampledinbonemarrowand/orperipheralbloodcellsofanimals,usuallyrodents.
2. Thepurposeofthemicronucleustestistoidentifysubstancesthatcausecytogenetic
damagewhichresultsintheformationofmicronucleicontaininglaggingchromosomefragmentsor
wholechromosomes.
3. Whenabonemarrowerythroblastdevelopsintoapolychromaticerythrocyte,themain
nucleusisextruded;anymicronucleusthathasbeenformedmayremainbehindintheotherwise
anucleatedcytoplasm.Visualisationofmicronucleiisfacilitatedinthesecellsbecausetheylacka
mainnucleus.Anincreaseinthefrequencyofmicronucleatedpolychromaticerythrocytesintreated
animalsisanindicationofinducedchromosomedamage.
4. DefinitionsusedaresetoutintheAnnex.
INITIALCONSIDERATIONS
5. Thebonemarrowofrodentsisroutinelyusedinthistestsincepolychromaticerythrocytes
areproducedinthattissue. Themeasurementofmicronucleatedimmature(polychromatic)
erythrocytesinperipheralbloodisequallyacceptableinanyspeciesinwhichtheinabilityofthe
spleentoremovemicronucleatederythrocyteshasbeendemonstrated,orwhichhasshownan
adequatesensitivitytodetectagentsthatcausestructuralornumericalchromosomeaberrations.
MpriecsreonncuecloeriacbasnenbceedoifstainkgiuniesthoecdhobryeoarncuenmtbreormeorficcrDitNerAia.inTthheesmeicirnocnluucdleei.idenTthieficfarteiqonuenocfythoef
micronucleatedimmature(polychromatic)erythrocytesistheprincipalendpoint. Thenumberof
mature(normochromatic)erythrocytesintheperipheralbloodthatcontainmicronucleiamonga
givennumberofmatureerythrocytescanalsobeusedastheendpointoftheassaywhenanimalsare
treatedcontinuouslyfor4weeksormore.
6. Thismammalianinvivomicronucleustestisespeciallyrelevanttoassessingmutagenic
hazardinthatitallowsconsiderationoffactorsofinvivometabolism,pharmacokineticsandDNA-
repairprocessesalthoughthesemayvaryamongspecies,amongtissuesandamonggenetic
endpoints.Aninvivoassayisalsousefulforfurtherinvestigationofamutageniceffectdetectedby
aninvitrosystem.
7. Ifthereisevidencethatthetestsubstance,orareactivemetabolite,willnotreachthetarget
tissue,itisnotappropriatetousethistest.
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PRINCIPLEOFTHETESTMETHOD
8. Animalsareexposedtothetestsubstancebyanappropriateroute.Ifbonemarrowisused,
theanimalsaresacrificedatappropriatetimesaftertreatment,thebonemarrowextracted,and
preparationsmadeandstained(1)(2)(3)(4)(5)(6)(7). Whenperipheralbloodisused,thebloodis
collectedatappropriatetimesaftertreatmentandsmearpreparationsaremadeandstained
(4)(8)(9)(10).Forstudieswithperipheralblood,aslittletimeaspossibleshouldelapsebetweenthe
lastexposureandcellharvest.Preparationsareanalyzedforthepresenceofmicronuclei.
DESCRIPTIONOFTHEMETHOD
Preparations
Selectionofanimalspecies
9. Miceorratsarerecommendedifbonemarrowisused,althoughanyappropriate
mammalianspeciesmaybeused.Whenperipheralbloodisused,micearerecommended.However,
anyappropriatemammalianspeciesmaybeusedprovideditisaspeciesinwhichthespleendoesnot
removemicronucleatederythrocytesoraspecieswhichhasshownanadequatesensitivitytodetect
agentsthatcausestructuralornumericalchromosomeaberrations. Commonlyusedlaboratory
strainsofyounghealthyanimalsshouldbeemployed. Atthecommencementofthestudy,the
weightvariationofanimalsshouldbeminimalandnotexceed±20%ofthemeanweightofeachsex.
Housingandfeedingconditions
10. Thetemperatureintheexperimentalanimalroomshouldbe22°C(±3°C). Althoughthe
relativehumidityshouldbeatleast30%andpreferablynotexceed70%otherthanduringroom
cleaning,theaimshouldbe50-60%.Lightingshouldbeartificial,thesequencebeing12hourslight,
12hoursdark.Forfeeding,conventionallaboratorydietsmaybeusedwithanunlimitedsupplyof
drinkingwater.Thechoiceofdietmaybeinfluencedbytheneedtoensureasuitableadmixtureofa
testsubstancewhenadministeredbythisroute.Animalsmaybehousedindividually,orcagedin
smallgroupsofthesamesex.
Preparationoftheanimals
11. Healthyyoungadultanimalsarerandomlyassignedtothecontrolandtreatmentgroups.
Theanimalsareidentifieduniquely. Theanimalsareacclimatedtothelaboratoryconditionsforat
leastfivedays.Cagesshouldbearrangedinsuchawaythatpossibleeffectsduetocageplacement
areminimized.
Preparationofdoses
12. Solidtestsubstancesshouldbedissolvedorsuspendedinappropriatesolventsorvehicles
anddiluted,ifappropriate,priortodosingoftheanimals. Liquidtestsubstancesmaybedosed
directlyordilutedpriortodosing. Freshpreparationsofthetestsubstanceshouldbeemployed
unlessstabilitydatademonstratetheacceptabilityofstorage.
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Testconditions
Solvent/vehicle
13. Thesolvent/vehicleshouldnotproducetoxiceffectsatthedoselevelsused,andshouldnot
besuspectedofchemicalreactionwiththetestsubstance.Ifotherthanwell-knownsolvents/vehicles
areused,theirinclusionshouldbesupportedwithreferencedataindicatingtheircompatibility.Itis
recommendedthatwhereverpossible,theuseofanaqueoussolvent/vehicleshouldbeconsidered
first.
Controls
14. Concurrentpositiveandnegative(solvent/vehicle)controlsshouldbeincludedforeachsex
ineachtest.Exceptfortreatmentwiththetestsubstance,animalsinthecontrolgroupsshouldbe
handledinanidenticalmannertoanimalsofthetreatmentgroups.
15. Positivecontrolsshouldproducemicronucleiinvivoatexposurelevelsexpectedtogivea
detectableincreaseoverbackground.Positivecontroldosesshouldbechosensothattheeffectsare
clearbutdonotimmediatelyrevealtheidentityofthecodedslidestothereader.Itisacceptablethat
thepositivecontrolbeadministeredbyaroutedifferentfromthetestsubstanceandsampledatonlya
singletime. Inaddition,theuseofchemicalclass-relatedpositivecontrolchemicalsmaybe
considered,whenavailable.Examplesofpositivecontrolsubstancesinclude:
ChemicalandCASNo.
Ethylmethanesulphonate[CASno.62-50-0]
Ethylnitrosourea[CASno.759-73-9]
MitomycinC[CASno.50-07-7]
Cyclophosphamide(monohydrate)[CASno.50-18-0(CASno.6055-19-2)]
Triethylenemelamine[CASno.51-18-3]
16. Negativecontrols,treatedwithsolventorvehiclealone,andotherwisetreatedinthesame
wayasthetreatmentgroupsshouldbeincludedforeverysamplingtime,unlessacceptableinter-
animalvariabilityandfrequenciesofcellswithmicronucleiaredemonstratedbyhistoricalcontrol
data. Ifsinglesamplingisappliedfornegativecontrols,themostappropriatetimeisthefirst
samplingtime. Inaddition,untreatedcontrolsshouldalsobeusedunlesstherearehistoricalor
publishedcontroldatademonstratingthatnodeleteriousormutageniceffectsareinducedbythe
chosensolvent/vehicle.
17. Ifperipheralbloodisused,apre-treatmentsamplemayalsobeacceptableasaconcurrent
negativecontrol,butonlyintheshortperipheralbloodstudies(e.g.,1-3treatment(s))whenthe
resultingdataareintheexpectedrangeforthehistoricalcontrol.
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PROCEDURE
N18u.mberandsexofanimals
Eachtreatedandcontrolgroupmustincludeatleast5analysableanimalspersex(11).If
atthetimeofthestudytherearedataavailablefromstudiesinthesamespeciesandusingthesame
routeofexposurethatdemonstratethattherearenosubstantialdifferencesbetweensexesintoxicity,
thentestinginasinglesexwillbesufficient. Wherehumanexposuretochemicalsmaybesex-
specific,asforexamplewithsomepharmaceuticalagents,thetestshouldbeperformedwithanimals
oftheappropriatesex.
Treatmentschedule
19. No standardtreatmentschedule(i.e.1,2,ormoretreatmentsat24hintervals)canbe
recommended.Thesamplesfromextendeddoseregimensareacceptableaslongasapositiveeffect
hasbeendemonstratedforthisstudyor,foranegativestudy,aslongastoxicityhasbeen
demonstratedorthelimitdosehasbeenused,anddosingcontinueduntilthetimeofsampling.Test
substancesmayalsobeadministeredasasplitdose,i.e.,twotreatmentsonthesamedayseparatedby
nomorethanafewhours,tofacilitateadministeringalargevolumeofmaterial.
20. Thetestmaybeperformedintwoways:
(a) Animalsaretreatedwiththetestsubstanceonce.Samplesofbonemarrowaretakenat
leasttwice,startingnotearlierthan24hoursaftertreatment,butnotextendingbeyond
48hoursaftertreatmentwithappropriateinterval(s)betweensamples. Theuseof
samplingtimesearlierthan24hoursaftertreatmentshouldbejustified. Samplesof
peripheralbloodaretakenatleasttwice,startingnotearlierthan36hoursafter
treatment,withappropriateintervalsfollowingthefirstsample,butnotextending
beyond72hours. Whenapositiveresponseisrecognizedatonesamplingtime,
additionalsamplingisnotrequired.
(b) If2ormoredailytreatmentsareused(e.g.twoormoretreatmentsat24hourintervals),
samplesshouldbecollectedoncebetween18and24hoursfollowingthefinal
treatmentforthebonemarrowandoncebetween36and48hoursfollowingthefinal
treatmentfortheperipheralblood(12).
21. Othersamplingtimesmaybeusedinaddition,whenrelevant.
Doselevels
22. Ifarangefindingstudyisperformedbecausetherearenosuitabledataavailable,itshould
beperformedinthesamelaboratory,usingthesamespecies,strain,sex,andtreatmentregimentobe
usedinthemainstudy(13).Ifthereistoxicity,threedoselevelsareusedforthefirstsamplingtime.
Thesedoselevelsshouldcoverarangefromthemaximumtolittleornotoxicity. Atthelater
samplingtimeonlythehighestdoseneedstobeused. Thehighestdoseisdefinedasthedose
producingsignsoftoxicitysuchthathigherdoselevels,basedonthesamedosingregimen,wouldbe
expectedtoproducelethality. Substanceswithspecificbiologicalactivitiesatlownon-toxicdoses
(suchashormonesandmitogens)maybeexceptionstothedose-settingcriteriaandshouldbe
evaluatedonacase-by-casebasis.Thehighestdosemayalsobedefinedasadosethatproduces
someindicationoftoxicityofthebonemarrow(e.g.areductionintheproportionofimmature
erythrocytesamongtotalerythrocytesinthebonemarroworperipheralblood).
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L23i.mittest
Ifatestatonedoselevelofatleast2000mg/kgbodyweightusingasingletreatment,oras
twotreatmentsonthesameday,producesnoobservabletoxiceffects,andifgenotoxicitywouldnot
beexpectedbasedupondatafromstructurallyrelatedsubstances,thenafullstudyusingthreedose
levelsmaynotbeconsiderednecessary. Forstudiesofalongerduration,thelimitdoseis2000
mg/kg/bodyweight/dayfortreatmentupto14days,and1000mg/kg/bodyweight/dayfortreatment
longerthan14days.Expectedhumanexposuremayindicatetheneedforahigherdoseleveltobe
usedinthelimittest.
Administrationofdoses
24. Thetestsubstanceisusuallyadministeredbygavageusingastomachtubeorasuitable
intubationcannula,orbyintraperitonealinjection. Otherroutesofexposuremaybeacceptable
wheretheycanbejustified.Themaximumvolumeofliquidthatcanbeadministeredbygavageor
injectionatonetimedependsonthesizeofthetestanimal. Thevolumeshouldnotexceed2
ml/lOOgbodyweight.Theuseofvolumeshigherthanthesemustbejustified.Exceptforirritating
orcorrosivesubstanceswhichwillnormallyrevealexacerbatedeffectswithhigherconcentrations,
variabilityintestvolumeshouldbeminimisedbyadjustingtheconcentrationtoensureaconstant
volumeatalldoselevels.
Bonemarrow/bloodpreparation
25. Bonemarrowcellsareusuallyobtainedfromthefemursortibiasimmediatelyfollowing
sacrifice.Commonly,cellsareremovedfromfemursortibias,preparedandstainedusing
establishedmethods. Peripheralbloodisobtainedfromthetailveinorotherappropriateblood
vaensdsetlh.enBsltoaoidnedc.ellTshearuesiemomfedaiaDtNelAysspteaciinfeidcssutparianvi[tea.gl.lyac(r8i)d(i9n)(e1o0r)aonrgesm(e14a)roprreHpoaercathisotns33a2re58mapldues
pyronin-Y(15)]caneliminatesomeoftheartifactsassociatedwithusinganon-DNAspecificstain.
Thisadvantagedoesnotprecludetheuseofconventionalstains(e.g.,Giemsa).Additionalsystems
[e.g.cellulosecolumnstoremovenucleatedcells(16)]canalsobeusedprovidedthatthesesystems
havebeenshowntoadequatelyworkformicronucleuspreparationinthelaboratory.
Analysis
26. Theproportionofimmatureamongtotal(immature+mature)erythrocytesisdetermined
foreachanimalbycountingatotalofatleast200erythrocytesforbonemarrowand1000
erythrocytesforperipheralblood(17).Allslides,includingthoseofpositiveandnegativecontrols,
shouldbeindependentlycodedbeforemicroscopicanalysis.Atleast2000immatureerythrocytesper
animalarescoredfortheincidenceofmicronucleatedimmatureerythrocytes. Additional
informationmaybeobtainedbyscoringmatureerythrocytesformicronuclei.Whenanalysingslides,
theproportionofimmatureerythrocytesamongtotalerythrocytesshouldnotbelessthan20%ofthe
controlvalue. Whenanimalsaretreatedcontinuouslyfor4weeksormore,atleast2000mature
erythrocytesperanimalcanalsobescoredfortheincidenceofmicronuclei.Systemsforautomated
analysis(imageanalysisandcellsuspensionsflowcytometry)areacceptablealternativestomanual
evaluationifappropriatelyjustifiedandvalidated.
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DATAANDREPORTING
Treatmentofresults
27. Individualanimaldatashouldbepresentedintabularform. Theexperimentalunitisthe
animal. Thenumberofimmatureerythrocytesscored,thenumberofmicronucleatedimmature
erythrocytes,andthenumberofimmatureamongtotalerythrocytesshouldbelistedseparatelyfor
eachanimalanalysed. Whenanimalsaretreatedcontinuouslyfor4weeksormore,thedataon
matureerythrocytesshouldalsobegivenifitiscollected.Theproportionofimmatureamongtotal
erythrocytesand,ifconsideredapplicable,thepercentageofmicronucleatederythrocytesisgivenfor
eachanimal.Ifthereisnoevidenceforadifferenceinresponsebetweenthesexes,thedatafrom
bothsexesmaybecombinedforstatisticalanalysis.
Evaluationandinterpretationofresults
28. Thereareseveralcriteriafordeterminingapositiveresult,suchasadose-relatedincreasein
thenumberofmicronucleatedcellsoraclearincreaseinthenumberofmicronucleatedcellsina
singledosegroupatasinglesamplingtime.Biologicalrelevanceoftheresultsshouldbeconsidered
first. Statisticalmethodsmaybeusedasanaidinevaluatingthetestresults(18)(19). Statistical
significanceshouldnotbetheonlydeterminingfactorforapositiveresponse. Equivocalresults
shouldbeclarifiedbyfurthertestingpreferablyusingamodificationofexperimentalconditions.
29. Atestsubstanceforwhichtheresultsdonotmeettheabovecriteriaisconsiderednon-
mutagenicinthistest.
30. Althoughmostexperimentswillgiveclearlypositiveornegativeresults,inrarecasesthe
datasetwillprecludemakingadefinitejudgementabouttheactivityofthetestsubstance.Results,
mayremainequivocalorquestionableregardlessofthenumberoftimestheexperimentisrepeated.
Positiveresultsinthemicronucleustestindicatethatasubstanceinducesmicronucleiwhicharethe
resultofchromosomaldamageordamagetothemitoticapparatusintheerythroblastsofthetest
species.Negativeresultsindicatethat,underthetestconditions,thetestsubstancedoesnotproduce
micronucleiintheimmatureerythrocytesofthetestspecies.
31. Thelikelihoodthatthetestsubstanceoritsmetabolitesreachthegeneralcirculation or
specificallythetargettissue (e.g.systemictoxicity)shouldbediscussed.
Testreport
32. Thetestreportshouldalsoincludethefollowinginformation:
Testsubstance:
- identificationdataandCASno.,ifknown;
physicalnatureandpurity;
- physiochemicalpropertiesrelevanttotheconductofthestudy;
stabilityofthetestsubstance,ifknown.
Solvent/vehicle:
- justificationforchoiceofvehicle;
solubilityandstabilityofthetestsubstanceinthesolvent/vehicle,ifknown.
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Testanimals:
species/strainused;
number,ageandsexofanimals;
source,housingconditions,diet,etc.;
individualweightoftheanimalsatthestartofthetest,includingbodyweightrange,
meanandstandarddeviationforeachgroup.
Testconditions:
positiveandnegative(vehicle/solvent)controldata;
datafromrange-findingstudy,ifconducted;
rationalefordoselevelselection;
- detailsoftestsubstancepreparation;
detailsoftheadministrationofthetestsubstance;
rationaleforrouteofadministration;
methodsforverifyingthatthetestsubstancereachedthegeneralcirculationortarget
tissue,ifapplicable;
conversionfromdiet/drinkingwatertestsubstanceconcentration(ppm)totheactual
dose(mg/kgbodyweight/day),ifapplicable;
detailsoffoodandwaterquality;
- detaileddescriptionoftreatmentandsamplingschedules;
methodsofslidepreparation;
methodsformeasurementoftoxicity;
criteriaforscoringmicronucleatedimmatureerythrocytes;
numberofcellsanalysedperanimal;
criteriaforconsideringstudiesaspositive,negativeorequivocal.
Results:
signsoftoxicity;
proportionofimmatureerythrocytesamongtotalerythrocytes;
numberofmicronucleatedimmatureerythrocytes,givenseparatelyforeachanimal;
mean+standarddeviationofmicronucleatedimmatureerythrocytespergroup;
dose-responserelationship,wherepossible;
statisticalanalysesandmethodapplied;
concurrentandhistoricalnegativecontroldata;
concurrentpositivecontroldata.
Discussionoftheresults.
Conclusion.
LITERATURE
(1) Heddle,J.A.(1973).ARapidInVivoTestforChromosomalDamage,MutationRes.,18,187-
190.
(2) Schmid,W.(1975).TheMicronucleusTest,MutationRes.,31,9-15.
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(3) Heddle,J.A.,Salamone,M.F.,Hite,M„Kirkhart,B„Mavoumin,K.,MacGregor,J.G.and
Newell,G.W.(1983).TheInductionofMicronucleiasaMeasureofGenotoxicity.Mutation
Res.123.61-118.
(4) MViavvoouMmicirno,nuKc.lH.e,usBlAaskseayy,Din.HM.a,mCmiamlinioa,nMB.oC.n,eSMaalrarmoonwe,anMd.FP.erainphdeHreadldBlleo,oJd..A.A(1r9e9p0o)r.tTohfetIhen
U.S.EnvironmentalProtectionAgencyGene-ToxProgram,MutationRes.,239,29-80.
(5) MErayctGhrreocgyotre,s:J.AT.,RSacphildegeSlc,reRe.nCfhoory,ChWr.oNm.o,saonmdalWeDharm,aCg.eM.Du(r1i9n83g).RoMuitcirnoenuTcolxeiiciitnyCiTrecsutliantginign
Mice,in:“DevelopmentsinScienceandPracticeofToxicology”,Ed.A.W.Hayes,R.C.
SchncllandT.S.Miya,Elsevier,Amsterdam,pp.555-558.
(6) MacGregor,J.T.,Heddle,J.A.,Hite,M„Margolin,G.H.,RamelC„Salamone,M.F.,Tice,R.R.
andWild,D.(1987).GuidelinesfortheConductofMicronucleusAssaysinMammalianBone
MarrowErythrocytes.MutationRes.,189,103-112.
(7) MacGregor,J.T.,Wehr,C.M.,Henika,P.R.,andShelby,M.E.(1990).TheinvivoErythrocyte
MicronucleusTest:MeasurementatSteadyStateIncreasesAssayEfficiencyandPermits
IntegrationwithToxicityStudies.Fund.Appl.Toxicol.,14,513-522.
(8) Hayashi,M.,Morita,T.,Kodama,Y.,Sofuni,T.,andIshidate,M.Jr.(1990).TheMicronucleus
AssaywithMousePeripheralBloodReticulocytesUsingAcridineOrange-CoatedSlides.
MutationResearch,245,245-249.
(9) TheCollaborativeStudyGroupfortheMicronucleusTest(1992).MicronucleusTestwith
MousePeripheralBloodErythrocytesbyAcridineOrangeSupravitalStaining:TheSummary
Reportofthe5thCollaborativeStudybyCSGMT/JEMS.MMS.MutationRes.,278,83-98.
(10) TheCollaborativeStudyGroupfortheMicronucleusTest(CSGMT/JEMMS.MMS,The
MammalianMutagenesisStudyGroupoftheEnvironmentalMutagenSocietyofJapan)(1995).
Protocol recommended forthe short-term mouse peripheral blood micronucleus test.
Mutagenesis,10,153-159.
(11) Hayashi,M„Tice,R.R.,MacGregor,J.T.,Anderson,D.,Blakey,D.H.,Kirsch-Volders,M„
Oleson,Jr.F.B..,Pacchierotti,F.,Romagna,F.,Shimada,H.,Sutou.S.andVannier,B.(1994).
InVivoRodentErythrocyteMicronucleusAssay.MutationRes.,312,293-304.
(12) Hdioguabslheikduonsii,ngN.inatnhdeSmuotuosue,pS.er(i1p9h9e5r)a.lAblnooodptmiimcarl,onguecnleeruaslitzeestd.sMaumtpalgienngestiism,e1o0f,33013+/--3169.hafter
(13) Fielder,R.J„Allen,J.A„Boobis,A.R„Botham,P.A.,Doe,J„Esdaile,D.J„Gatehouse,D.
G.,Hodson-Walker,G.,Morton,D.B„Kirkland,D.J.,andRichold,M.(1992).Reportof
BritishToxicologySociety/UKEnvironmentalMutagenSocietyWorkingGroup:DoseSetting
inInVivoMutagenicityAssays.Mutagenesis,7,313-319.
(14) Hayashi,M.,Sofuni,T.,andIshidate,M.Jr.(1983).AnApplicationofAcridineOrange
FluorescentStainingtotheMicronucleusTest.MutationRes.,120.241-247.
(15) MPraoccGerdeugroer,foJr.MTi„crWoenuhcrl,eiC.anMd„RaNnAdLianngElroyitsh,roRc.ytGe.s(U1s9i8n3g).HAoecShismtpl3e3F2l5u8oraensdcenPtyrSotnaiinniYn.g
MutationRes.,120,269-275.
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(16) Romagna,F.andStaniforth,C.D.(1989).Theautomatedbonemarrowmicronucleustest.
MutationRes.,213,91-104.
(17) Gollapudi,B.andMcFadden,L.G.(1995).Samplesizefortheestimationofpolychromatic
tonormochromaticeruthrocyteratiointhebonemarrowmicronucleustest.MutationRes.,
347.97-99.
(18) IRRniecchVooilmvdmo,eMnC.yd,teoAdgsehnPberyot.ciecJds.u,rBAeossso.atymUsa,Kn,EInMJ:.,SCDhJSa.unKbdiclroekmylm,ainAtd.t,eG(eEadto.en)hoGBuuasisedi,eclDi.MnGue.tsaagfneodrniMHcueittnaydgeerTnseiosctnis.t,yL.UT(Ke1sE9t9iM0n)gS..
Report.PartIrevised.CambridgeUniversityPress,Cambridge,NewYork,PortChester,
Melbourne,Sydney,pp.115-141.
(19) MALos„vsealPylas,pIwDn.o:Pr.Dt,.hJ..AnDKd.ierGrk.slaoannnd,d(DSE„da.v)AalgSbeta,antieJss.teRi,.caKlR.„E(v1a9Al8mu9pa)ht.lieoStnttao,tfisGMt.iuEct.aa,lgeCAnlnaiarcleiy,tsiyGs.T,eosFfterIDngautsVaoi.nv,oUKRC.yE,tMoRgiSecnhSeoultbdi,-c
committeeonGuidelinesforMutagenicityTesting,Report,PartHI.CambridgeUniversity
Press,Cambridge,NewYork,PortChester,Melbourne,Sydney,pp.184-232.
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ANNEX
DEFINITIONS
Cceelnltdriovmiseiroen,(aKlilnoewtionchgoorred)e:rlRyegmioovn(esm)enoftaofcdharuogmhotseromcehrwoimtohswohmiecshtsoptihnedlpeolfeibseorsftahreedaasusgochitaetrecdeldlusr.ing
Mteilcorpohnausceleoif:mistmoaslilsn(umcelieois,iss)epbayraltaeggfirnogmcharnodmaodsdoitmieonfarlagtometnhtesmoraiwnhonlueclcehiroofmocselolms,esp.roducedduring
Nimomramtoucrher,opmoaltyicchreormyatthircoceyrtyet:hrmoactyutreesberyysttharioncsytseeletchtaitvelafcokrsrribiobsoosmoemse.sandcanbedistinguishedfrom
Pcoonltyacihnrsomraitbiocsoemreystharnocdytteh:eriemfmoraetucraenebreytdhirsotciyntgeu,isihnedanfrionmtermmaetduiraet,ensotramgoecohfrodmeavteilcopemreyntth,roctyhtatesstbilyl
stainsselectiveforribosomes.
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