natgillin/translations-raw huggingface dataset:
b. Benyttes der to daglige behandlinger (f.eks. to behandlinger med to timers mellemrum) udtages der for knoglemarvs vedkommende prøver en gang mellem 18 og 24 timer efter den sidste behandling, eller for perifert blods vedkommende en gang mellem 36 og 48 timer efter den sidste behandling (30).
Mice, rats, or another appropriate mammalian species may be used.
c. Benyttes der tre eller flere daglige behandlinger (f.eks. tre eller flere behandlinger med ca. 24 timers mellemrum), bør knoglemarvsprøver indsamles senest 24 timer efter den sidste behandling, og perifert blod bør indsamles senest 40 timer efter den sidste behandling (31).
When peripheral blood is used, it must be established that splenic removal of micronucleated cells from the circulation does not compromise the detection of induced micronuclei in the species selected.
Denne behandlingsmulighed gør det muligt at kombinere kometassays (f.eks. prøveudtagning 2-6 timer efter den sidste behandling) med mikrokernetesten og integrere mikrokernetesten med undersøgelserne af toksicitet ved gentagen indgivelse.
This has been clearly demonstrated for mouse and rat peripheral blood (2).
Akkumulerede data tydede på, at der kan observeres mikrokerneinduktion i disse bredere tidsrammer ved tre eller flere indgivelser (15).
If species other than rodents are used, it is recommended that the measurement of induced micronuclei be integrated into another appropriate toxicity test.
Andre doserings- eller prøveudtagningsregimer kan anvendes, hvor det er relevant og videnskabeligt begrundet og for at fremme integrationen med andre toksicitetstest.
Test Conditions
Alle dyr bør vejes ved undersøgelsens start, mindst en gang om ugen i undersøgelser med gentagne doser og ved aflivning.
The solvent/vehicle should not produce toxic effects at the dose levels used, and should not be capable of chemical reaction with the test chemicals.
Dyr, der udviser ikkedødelig indikation for for høj toksicitet, bør aflives humant før testperiodens udløb (28).
In the absence of historical or published control data showing that no micronuclei and other deleterious effects are induced by a chosen atypical solvent/vehicle, an initial study should be conducted in order to establish the acceptability of the solvent/vehicle control.
Under visse omstændigheder kan dyrs kropstemperatur overvåges, da behandlingsinduceret hyper- og hypotermi har været medvirkende til at frembringe falske resultater (32) (33) (34).
When a concurrent positive control group is not included, scoring controls (fixed and unstained slides or cell suspension samples, as appropriate for the method of scoring) should be included in each experiment.
Der bør på passende tidspunkt(er) tages en blodprøve for at gøre det muligt at undersøge plasmaniveauet i testkemikaliet med henblik på at påvise, at der er sket eksponering af knoglemarven, hvor der er grundlag for dette, og hvor der ikke findes andre eksponeringsdata (jf. punkt 48).
These can be obtained by including within the scoring of the study appropriate reference samples that have been obtained and stored from a separate positive control experiment conducted periodically (e.g. every 6-18 months); for example, during proficiency testing and on a regular basis thereafter, where necessary.
Præparering af knoglemarv/blod
Positive control chemicals should reliably produce a detectable increase in micronucleus frequency over the spontaneous level.
Knoglemarvceller udtages normalt fra dyrenes lårben eller skinneben umiddelbart efter human aflivning.
When employing manual scoring by microscopy, positive control doses should be chosen so that the effects are clear but do not immediately reveal the identity of the coded samples to the scorer.
Sædvanligvis udtages cellerne, hvorefter de præpareres og farves med velkendte metoder.
Chemicals and CASRN
Der kan udtages små mængder perifert blod i henhold til passende standarder for dyrevelfærd, enten ved hjælp af en metode, der lader forsøgsdyret overleve, som f.eks. ved blødning fra halevenen eller et andet egnet blodkar, eller ved hjertepunktur eller prøveudtagning fra et stort blodkar ved aflivning.
Ethyl methanesulphonate [CASRN 62-50-0]
For både knoglemarv eller erythrocytter fra perifert blod kan cellerne, afhængigt af analysemetoden, straks farves supravitalt (16) (17) (18), idet udstrygningspræparater fremstilles og derefter farves til mikroskopi eller fikseres og farves behørigt med henblik på flowcytometrianalyse.
Methyl methanesulphonate [CASRN 66-27-3]
Ved at bruge en DNA-specifik farvning [f.eks. acridinorange (35) eller Hoechst 33258 plus pyronin-Y (36)] kan man undgå nogle af de artefakter, der optræder ved ikke-DNA-specifik farvning.
Ethyl nitrosourea [CASRN 759-73-9]
Denne fordel udelukker ikke, at der benyttes konventionelle farvestoffer (f.eks. Giemsa ved mikroskopianalyse).
Mitomycin C [CASRN 50-07-7]
Supplerende systemer [f.eks. cellulosekolonner til fjernelse af kerneindeholdende celler (37) (38)] kan også benyttes, forudsat at disse systemer påviseligt er kompatible med præpareringen af prøven i laboratoriet.
Cyclophosphamide (monohydrate) [CASRN 50-18-0 (CASRN 6055-19-2)]
Hvis disse metoder kan anvendes, kan der anvendes anti-kinetokor-antistoffer (39), FISH med pancentromeriske DNA-prober (40) eller primet in situ-mærkning med pancentromerspecifikke primere og egnet DNA-kontrastfarvning (41) til at identificere arten af mikrokerner (kromosom/kromosomfragment) med henblik på at fastslå, om induktionen af mikrokerner skyldes klastogen og/eller aneugen aktivitet.
Triethylenemelamine [CASRN 51-18-3]
Andre metoder til differentiering mellem klastogener og aneugener kan anvendes, hvis de har vist sig at være effektive.
Colchicine [CASRN 64-86-8] or Vinblastine [CASRN 865-21-4] — as aneugens
Analyse (manuel og automatisk)
However, if consistent inter-animal variability and frequencies of cells with micronuclei are demonstrated by historical negative control data at each sampling time for the testing laboratory, only a single sampling for the negative control may be necessary.
Alle objektglas eller prøver til analyse, herunder til positive og negative kontroller, bør kodes uafhængigt inden nogen form for analyse og bør randomiseres, så den manuelle bedømmer ikke har kendskab til behandlingsbetingelsen; denne bedømmelse er ikke nødvendig, når der anvendes automatiske scoringssystemer, som ikke bygger på visuel inspektion og ikke kan påvirkes af operatørbias.
If peripheral blood is used, a pre-treatment sample is acceptable instead of a concurrent negative control for short-term studies when the resulting data are consistent with the historical control database for the testing laboratory.
For hvert dyr bestemmes andelen af umodne erythrocytter i forhold til det samlede antal (umodne og modne) erythrocytter, idet der tælles mindst 500 erythrocytter i alt for knoglemarv og 2000 erythrocytter for perifert blod (42).
It has been shown for rats that pre-treatment sampling of small volumes (e.g. below 100 μl/day) has minimal impact on micronucleus background frequency (25).
For hvert dyr scores mindst 4000 umodne erythrocytter for forekomsten af umodne etythrocytter med mikrokerner (43).
In general, the micronucleus response is similar between male and female animals and, therefore, most studies could be performed in either sex (26).
Hvis den historiske negative kontroldatabase viser, at den gennemsnitlige baggrundsfrekvens for umodne erythrocytter med mikrokerner er < 0,1 % i testlaboratoriet, bør det overvejes at score flere celler.
Data demonstrating relevant differences between males and females (e.g. differences in systemic toxicity, metabolism, bioavailability, bone marrow toxicity, etc. including e.g. in a range-finding study) would encourage the use of both sexes.
Når prøverne analyseres, bør andelen af umodne erythrocytter i forhold til det samlede antal erythrocytter hos behandlede dyr ikke være mindre end 20 % af forholdet mellem bærestof-/opløsningsmiddelkontrol ved scoring ved mikroskopi og ikke mindre end ca. 5 % af forholdet mellem bærestof-/opløsningsmiddelkontrol ved scoring af CD71+ umodne erythrocytter CD71+ ved cytometrimetoder (jf. punkt 31) (29).
As a guide to maximum typical animal requirements, a study in bone marrow conducted according to the parameters established in paragraph 37 with three dose groups and concurrent negative and positive controls (each group composed of five animals of a single sex) would require between 25 and 35 animals.
For et knoglemarvsassay scoret ved mikroskopi gælder det f.eks., at hvis kontrolandelen af umodne erythrocytter i knoglemarven er 50 %, vil den øvre grænse for toksicitet være 10 % umodne erythrocytter.
If a preliminary range-finding study is performed because there are no suitable data already available to aid in dose selection, it should be performed in the same laboratory, using the same species, strain, sex, and treatment regimen to be used in the main study (27).
Da rottemilten udskiller og destruerer erythrocytter med mikrokerner, foretrækkes det for at fastholde en høj følsomhed i assayet ved analyse af perifert blod at begrænse analysen af umodne erythrocytter med mikrokerner til den yngste fraktion.
The study should aim to identify the maximum tolerated dose (MTD), defined as the highest dose that will be tolerated without evidence of study-limiting toxicity, relative to the duration of the study period (for example, by inducing body weight depression or hematopoietic system cytotoxicity, but not death or evidence of pain, suffering or distress necessitating humane euthanasia (28)).
Ved hjælp af automatiserede analysemetoder kan disse mest umodne erythrocytter identificeres på basis af deres høje indhold af RNA eller det høje niveau af transferrinreceptorer (CD71+) udtrykt på deres overflade (31).
The highest dose may also be defined as a dose that produces toxicity in the bone marrow (e.g. a reduction in the proportion of immature erythrocytes among total erythrocytes in the bone marrow or peripheral blood of more than 50 %, but to not less than 20 % of the control value).
En direkte sammenligning af forskellige farvningsmetoder har imidlertid vist, at der kan opnås et tilfredsstillende resultat med forskellige metoder, herunder traditionel acridinorangefarvning (3) (4).
However, when analysing CD71-positive cells in peripheral blood circulation (i.e., by flow cytometry), this very young fraction of immature erythrocytes responds to toxic challenges more quickly than the larger RNA-positive cohort of immature erythrocytes.
For hvert dyr registreres antallet af scorede umodne erythrocytter, antallet af umodne erythrocytter med mikrokerne og andelen af umodne erythrocytter i forhold til det samlede antal erythrocytter.
Therefore, higher apparent toxicity may be evident with acute exposure designs examining the CD71-positive immature erythrocyte fraction as compared to those that identify immature erythrocytes based on RNA content.
Når mus behandles kontinuerligt i fire uger eller derover, bør der også anføres data om antallet og andelen af modne erythrocytter med mikrokerner, hvis de indsamles.
For this reason, when experiments utilise five or fewer days of treatment, the highest dose level for test chemicals causing toxicity may be defined as the dose that causes a statistically significant reduction in the proportion of CD71-positive immature erythrocytes among total erythrocytes but not to less than 5 % of the control value (29).
a. De sideløbende negative kontroldata anses for at kunne føjes til laboratoriets historiske negative kontroldatabase (jf. punkt 15-18)).
Chemicals that exhibit saturation of toxicokinetic properties, or induce detoxification processes that may lead to a decrease in exposure after long-term administration may be exceptions to the dose-setting criteria and should be evaluated on a case-by-case basis.
b. Sideløbende positive kontroller eller scoringskontroller bør fremkalde responser, der er forenelige med responserne i den historiske positive kontroldatabase, og medføre en statistisk set signifikant stigning sammenlignet med den sideløbende negative kontrol (jf. punkt 24-25).
If the test chemical does not produce toxicity in a range-finding study or based on existing data, the highest dose for an administration period of 14 days or more should be 1000 mg/kg body weight/day, or for administration periods of less than 14 days, 2000 mg/kg/body weight/day.
c. Der er analyseret et passende antal doser og celler.
When administration occurs for 14 days or more, the limit dose is 1000 mg/kg body weight/day.
d. Kriterierne for valg af højeste dosis er i overensstemmelse med kriterierne i punkt 30-33.
For administration periods of less than 14 days, the limit dose is 2000 mg/kg/body weight/day.
a. Mindst en af behandlingsgrupperne viser en statistisk signifikant stigning i frekvensen af umodne erythrocytter med mikrokerner sammenlignet med den sideløbende negative kontrol.
Therefore, routes of exposure such as dietary, drinking water, topical subcutaneous, intravenous, oral (by gavage), inhalation, intratracheal, or implantation may be chosen as justified.
Anbefalinger til de mest hensigtsmæssige statistiske metoder kan findes i litteraturen (44) (45) (46) (47).
If the test chemical is admixed in diet or drinking water, especially in case of single dosing, care should be taken that the delay between food and water consumption and sampling should be sufficient to allow detection of the effects (see paragraph 37).
Positive resultater af mikrokernetesten viser, at testkemikaliet fremkalder mikrokerner, som er en følge af kromosombeskadigelse eller beskadigelse af mitoseapparatet hos erythroblaster i testarten.
Preferably, 2 or more treatments are performed, administered at 24-hour intervals, especially when integrating this test into other toxicity studies.
I det tilfælde, hvor der blev foretaget en test med henblik på at påvise centromerer i mikrokerner, er et testkemikalie, der producerer mikrokerner indeholdende centromer (DNA-centromer eller kinetokor, tegn på hele kromosomtab) tegn på, at testkemikaliet er et aneugen.
In the alternative, single treatments can be administered, if scientifically justified (e.g. test chemicals known to block cell cycle).
Såfremt alle acceptkriterier er opfyldt, anses et testkemikalie for at være klart negativt, hvis følgende er opfyldt under alle de undersøgte forsøgsbetingelser:
Test chemicals also may be administered as a split dose, i.e., two or more treatments on the same day separated by no more than 2-3 hours, to facilitate administering a large volume.
a. Ingen af behandlingsgrupperne viser en statistisk signifikant stigning i frekvensen af umodne erythrocytter med mikrokerner sammenlignet med den sideløbende negative kontrol.
The test may be performed in mice or rats in one of three ways:
Dokumentation for knoglemarvens eksponering for et testkemikalie kan omfatte et fald i forholdet mellem umodne og modne erythrocytter eller måling af plasma eller blodniveauer for testkemikaliet.
Animals are treated with the test chemical once.
I tilfælde af intravenøs indgivelse er der ikke behov for dokumentation for eksponering.
Samples of bone marrow are taken at least twice (from independent groups of animals), starting not earlier than 24 hours after treatment, but not extending beyond 48 hours after treatment with appropriate interval(s) between samples, unless a test chemical is known to have an exceptionally long half-life.
Negative resultater viser, at testkemikaliet under testbetingelserne ikke fremkalder mikrokerner hos umodne erythrocytter hos testarterne.
The use of sampling times earlier than 24 hours after treatment should be justified.
oprindelse, batchnummer, frist for anvendelse, hvis dette foreligger
Samples of peripheral blood are taken at least twice (from the same group of animals), starting not earlier than 36 hours after treatment, with appropriate interval(s) following the first sample, but not extending beyond 72 hours.
testkemikaliets opløselighed og stabilitet i opløsningsmidlet/bærestoffet, hvis dette er kendt
When a positive response is detected at one sampling time, additional sampling is not required unless quantitative dose-response information is needed.
positive og negative (bærestof/opløsningsmiddel) kontroldata
The described harvest times are a consequence of the kinetics of appearance and disappearance of the micronuclei in these 2 tissue compartments.
metoder til kontrol af, at testkemikaliet eller testkemikalierne er nået frem til det almindelige kredsløb eller målvævet
If 2 daily treatments are used (e.g. two treatments at 24 hour intervals), samples should be collected once between 18 and 24 hours following the final treatment for the bone marrow or once between 36 and 48 hours following the final treatment for peripheral blood (30).
kriterier for scoring af umodne erythrocytter med mikrokerner
If three or more daily treatments are used (e.g. three or more treatments at approximately 24 hour intervals), bone marrow samples should be collected no later than 24 hours after the last treatment and peripheral blood should be collected no later than 40 hours after the last treatment (31).
antal analyserede celler pr. dyr ved fastsættelse af frekvensen af umodne erythrocytter med mikrokerner og ved fastsættelse af forholdet mellem umodne og modne erythrocytter
This treatment option accommodates combination of the comet assay (e.g. sampling 2-6 hours after the last treatment) with the micronucleus test, and integration of the micronucleus test with repeated-dose toxicity studies.
metoder som f.eks. anvendelse af anti-kinetokor-antistoffer eller centromerspecifikke DNA-prober til bestemmelse af, om mikrokerner indeholder hele kromosomer eller kromosomfragmenter, hvor dette er relevant.
Accumulated data suggested that micronucleus induction can be observed over these wider timeframes when 3 or more administrations have occurred (15).
andel af umodne erythrocytter i forhold til samlet antal erythrocytter
Other dosing or sampling regimens may be used when relevant and scientifically justified, and to facilitate integration with other toxicity tests.
antal umodne erythrocytter med mikrokerner, anført særskilt for hvert dyr
All animals should be weighed at study initiation, at least once a week during repeated dose studies, and at euthanasia.
gennemsnit ± standardafvigelse for umodne erythrocytter med mikrokerner for hver gruppe
Animals exhibiting non-lethal indicators of excessive toxicity should be humanely euthanised prior to completion of the test period (28).
statistiske analyser og anvendte metoder
Under certain circumstances, animal body temperature could be monitored, since treatment-induced hyper- and hypothermia have been implicated in producing spurious results (32) (33) (34).
sideløbende negative og positive kontroldata med angivelse af intervaller, gennemsnit og standardafvigelser
A blood sample should be taken at appropriate time(s) in order to permit investigation of the plasma levels of the test chemicals for the purposes of demonstrating that exposure of the bone marrow occurred, where warranted and where other exposure data do not exist (see paragraph 48).
historisk negative og positive kontroldata med angivelse af intervaller, gennemsnit og standardafvigelser og kontrolgrænser på 95 % for fordelingen samt tidsperiode og antal datapunkter
Bone marrow / blood preparation
karakteriseringsdata, der viser, om mikrokerner indeholder hele kromosomer eller kromosomfragmenter, hvor dette er relevant
Bone marrow cells are usually obtained from the femurs or tibias of the animals immediately following humane euthanasia.
opfyldte kriterier for en positiv eller negativ respons.
Commonly, cells are removed, prepared and stained using established methods.
Higashikuni, N., S. Sutou (1995), An optimal, generalized sampling time of 30 +/- 6 h after double dosing in the mouse peripheral blood micronucleus test, Mutagenesis, Vol. 10/4, pp.
Small volumes of peripheral blood can be obtained, according to adequate animal welfare standards, either using a method that permits survival of the test animal, such as bleeding from the tail vein or other appropriate blood vessel, or by cardiac puncture or sampling from a large vessel at animal euthanasia.
313-319.
For both bone marrow or peripheral blood-derived erythrocytes, depending on the method of analysis, cells may be immediately stained supravitally (16) (17) (18), smear preparations are made and then stained for microscopy, or fixed and stained appropriately for flow cytometric analysis.
UKEMS SubCommittee on Guidelines for Mutagenicity Testing, Report, Part III, Kirkland, D.J. (ed.), Cambridge University Press, Cambridge, pp.
The use of a DNA specific stain [e.g. acridine orange (35) or Hoechst 33258 plus pyronin-Y (36)] can eliminate some of the artifacts associated with using a non-DNA specific stain.
Et tidligt stadium af erythrocytudvikling umiddelbart forud for den umodne erythrocyt, hvor cellen stadig indeholder en kerne.
This advantage does not preclude the use of conventional stains (e.g. Giemsa for microscopic analysis).
Kinetokor
Additional systems [e.g. cellulose columns to remove nucleated cells (37) (38)] also can be used provided that these systems have been demonstrated to be compatible with sample preparation in the laboratory.
Proteinstruktur, der dannes på centromeren af eukaryote celler, der forbinder kromosomet til mikrotubule polymerer fra mitotisk spindel under mitose og meiose og funktioner under celledeling til at trække søsterkromatider fra hinanden.
Where these methods are applicable, anti-kinetochore antibodies (39), FISH with pancentromeric DNA probes (40), or primed in situ labelling with pancentromere-specific primers, together with appropriate DNA counterstaining (41), can be used to identify the nature of the micronuclei (chromosome/chromosomal fragment) in order to determine whether the mechanism of micronucleus induction is due to clastogenic and/or aneugenic activity.
Mikrokerner
Other methods for differentiation between clastogens and aneugens may be used if they have been shown to be effective.
Små ekstrakerner, der er adskilt fra cellens hovedkerne, og som dannes under mitosens telofase (meiose) ud fra efterladte kromosomfragmenter eller hele kromosomer.
Analysis (manual and automated)
Normokromatisk eller moden erythrocyt
All slides or samples for analysis, including those of positive and negative controls, should be independently coded before any type of analysis and should be randomised so the manual scorer is unaware of the treatment condition; such coding is not necessary when using automated scoring systems which do not rely on visual inspection and cannot be affected by operator bias.
En fuldmoden erythrocyt, der har mistet den resterende RNA, der er tilbage efter enukleation, og/eller har mistet andre kortvarige cellemarkører, der typisk forsvinder efter enukleation efter den endelige erythroblastdeling.
The proportion of immature among total (immature + mature) erythrocytes is determined for each animal by counting a total of at least 500 erythrocytes for bone marrow and 2000 erythrocytes for peripheral blood (42).
Polykromatisk eller umoden erythrocyt
At least 4000 immature erythrocytes per animal should be scored for the incidence of micronucleated immature erythrocytes (43).
En nydannet erythrocyt på et udviklingsmellemstadium, som farves med både blå og røde elementer af klassiske blodfarver såsom Wrights Giemsa på grund af tilstedeværelsen af resterende RNA i den nydannede celle.
If the historical negative control database indicates the mean background micronucleated immature erythrocyte frequency is < 0,1 % in the testing laboratory, consideration should be given to scoring additional cells.
Disse nydannede celler er omtrent de samme som retikulocytter, som vises ved hjælp af en vital farvning, der får den resterende RNA til at klumpe sammen i et retikulum.
When analysing samples, the proportion of immature erythrocytes to total erythrocytes in treated animals should not be less than 20 % of the vehicle/solvent control proportion when scoring by microscopy and not less than approximately 5 % of the vehicle/solvent control proportion when scoring CD71+ immature erythrocytes by cytometric methods (see paragraph 31) (29).