natgillin/translations-raw huggingface dataset:
Når koncentrationerne ikke holder sig inden for 80-120 % af den nominelle koncentration, bør effektkoncentrationerne udtrykkes i forhold til den geometriske middelværdi af de målte koncentrationer, se kapitel 5 i OECD's vejledning: Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures (28).
Numerical Rating
GRÆNSETEST
Date
Med den i denne metode beskrevne fremgangsmåde kan der udføres en grænsetest ved 100 mg/l testkemikalie, dog ikke højere end kemikaliets opløselighed i testmediet, med henblik på at godtgøre, at LC50 er større end denne koncentration.
resent
Grænsetesten bør udføres med 20 embryoner i behandlingen, den positive kontrol og — om nødvendigt — opløsningsmiddelkontrollen og 24 embryoner i den negative kontrol.
Total Score
Hvis den procentvise dødelighed ved den testede koncentration overstiger dødeligheden i den negative kontrol (eller opløsningsmiddelkontrollen) med 10 %, bør der udføres en komplet test.
nlarged
Observerede virkninger bør registreres.
ronounced
Hvis dødeligheden overstiger 10 % i den negative kontrol (eller opløsningsmiddelkontrollen), bliver testen ugyldig og bør gentages.
Appendix 5B
I denne test betragtes de enkelte brønde som uafhængige paralleller til statistisk analyse.
ASSESSMENT OF SECONDARY SEX CHARACTERISTICS IN MEDAKA FOR THE DETECTION OF CERTAIN ENDOCRINE ACTIVE CHEMICALS
Der optegnes en kurve over procentdel embryoner, hvor mindst en af de apikale observationer er positiv ved 48 og/eller 96 timer, i forhold til testkoncentrationer.
Below is a description of the measurement of papillary processes [1], which are the secondary sex characteristics in medaka (Oryzias latipes).
Til beregning af kurvens hældning, LC50- værdier og konfidensgrænser (95 %) bør der anvendes passende statistiske metoder (38), og OECD's vejledning: Guidance Document on Current Approaches in the Statistical Analysis of Ecotoxicity Data bør konsulteres (39).
After the excision of the liver (Appendix 6), the carcass is placed into a conical tube containing about 10 ml of 10 % neutral buffered formalin (upside: head, downside: tail).
kemisk identifikation, f.eks. IUPAC-navn eller CAS-navn, CAS-nummer, SMILES- eller InChI-kode, strukturformel, renhed, kemisk identitet af urenheder i det omfang, det er relevant og praktisk muligt, osv. (herunder evt. organisk kulstofindhold).
If the gonad is fixed in a solution other than 10 % neutral buffered formalin, make a transverse cut across the carcass between anterior region of anal fin and anus using razor, taking care not to harm the gonopore and gonad itself (Fig.3).
Testorganismer:
Place the cranial side of the fish body into the fixative solution to preserve the gonad, and the tail side of the fish body into the 10 % neutral buffered formalin as described above.
testprocedure (f.eks. semistatisk udskiftning)
After placing the fish body into 10 % neutral buffered formalin, grasp the anterior region of the anal fin with tweezers and fold it for about 30 seconds to keep the anal fin open.
lysperiode
When grasping the anal fin with tweezers, grasp a few fin rays in the anterior region with care not to scratch the papillary processes.
testdesign (f.eks. antal testkamre, typer kontrol)
After keeping the anal fin open for about 30 seconds, store the fish body in 10 % neutral buffered formalin at room temperature until the measurement of the papillary processes (measurement should be conducted after fixing for at least 24 hours).
kvalitetskarakteristika for vandet ved hold af fisk (f.eks. pH, hårdhedsgrad, temperatur, ledningsevne og opløst ilt)
Measurement
koncentration af opløst ilt, pH, samlet hårdhedsgrad, temperatur og ledningsevne for testopløsningerne i starten og efter 96 timer
After fixing the fish body in the 10 % neutral buffered formalin for at least 24 hours, pick up the fish carcass from the conical tube and wipe the formalin on the filter paper (or paper towel).
metode til fremstilling af stamopløsninger og testopløsninger samt hyppighed af udskiftninger
Place the fish abdomen side up.
begrundelse for brug af bærestof og begrundelse for valg af opløsningsmiddel, hvis dette ikke er vand
Then cut the anal fin using small dissection scissors carefully (it is preferable to cut the anal fin with small amount of pterygiophore).
de nominelle testkoncentrationer og resultatet af alle analyser til bestemmelse af koncentrationen af testkemikaliet i testbeholderne; metodens genfindingsgrad og kvantificeringsgrænsen (LoQ) bør også rapporteres
Grasp the anterior region of the severed anal fin with tweezers and put it on a glass slide with a several drops of water.
bevis på, at kontrollerne opfyldte de generelle validitetskriterier for overlevelse
Then cover the anal fin with a cover glass.
æggenes befrugtningshastighed
Be careful not to scratch the papillary processes when grasping the anal fin with tweezers.
klækningshastighed i behandlings- og kontrolgrupper
Count the number of the joint plate with papillary processes using the counter under a biological microscope (upright microscope or inverted microscope).
højeste koncentration, der ikke forårsager dødelighed inden for testperioden
The papillary processes are recognised when a small formation of processes is visible on the posterior margin of joint plate.
mindste koncentration, der forårsager 100 % dødelighed inden for testperioden
Write the number of joint plate with papillary processes in each fin ray to the worksheet (e.g. first fin ray: 0, second fin ray: 10, third fin ray: 12, etc.) and enter the sum of this number on the Excel spreadsheet by individual fish.
kumulativ dødelighed for hver koncentration på de anbefalede observationstidspunkter
If necessary, take a photograph of the anal fin and count the number of joint plate with papillary processes on the photograph.
værdier efter 96 timer (og eventuelt efter 48 timer) for dødelighed med 95 %-konfidensgrænser, hvis det er muligt
After the measurement, put the anal fin into the conical tube described in (1) and store it.
afbildning af koncentration/dødelighedskurve ved testens afslutning
Fig.1.
dødelighed i kontrollerne (negative kontroller, interne pladekontroller samt positiv kontrol og anvendte opløsningsmiddelkontroller)
Diagram showing sexual difference in shape and size of the anal fin.
oplysninger om resultatet af hver af de fire apikale observationer
A, male; B, female.
forekomst og beskrivelse af eventuelle morfologiske og fysiologiske abnormiteter (jf. eksemplerne i tillæg 5, figur 2)
Fig.2.
hændelser i løbet af testen, der kan have påvirket resultaterne
A, Processes on joint plates of anal fin-ray.
statistisk analyse og behandling af data (probabalistiske analyser, model for logistisk regression og geometrisk middelværdi for LC50)
J.P., joint plate; A.S., axial space; P., process.
hældning og konfidensgrænser for regression af den (transformerede) koncentration/respons-kurve.
B, Distal extremity of fin-ray.
Afvigelser fra forsøgsmetoden og relevante forklaringer.
Actinotrichia (Act.) are on the tip.
Diskussion og fortolkning af resultater.
Fig.3.
OECD (2011) Validation Report (Phase 1) for the Zebrafish Embryo Toxicity Test: Part I and Part II.
Photograph of fish body showing the cut site when the gonad is fixed in the fixing solution other than 10 % neutral buffered formalin.
OECD (2012) Validation Report (Phase 2) for the Zebrafish Embryo Toxicity Test: Part I and Part II (Annexes).
In that case, the remaining body will be cut off between anterior region of anal fin and anal using razor (red bar), and the head side of fish body will be put into the fixing solution for gonad and the tail side of the fish body will be put into the 10 % neutral buffered formalin.
Environmental Toxicology and Chemistry, 32: 1768-1783.
Papillary processes normally appear only in adult males and are found on fin rays from the second to the seventh or eighth counting from the posterior end of the anal fin (Fig.1 and 2).
Pharmacol.: 149 (2), 196-209.
However, processes rarely appear on the first fin ray from the posterior end of the anal fin.
Chapter C.4 of this Annex: Ready Biodegradability.
This SOP covers the measurement of processes on the first fin ray (the fin ray number refers to the order from the posterior end of the anal fin in this SOP).
Chapter C.29 of this Annex: Ready Biodegradability, CO2 in sealed vessels.
RECOMMENDED PROCEDURES FOR SAMPLE COLLECTION FOR VITELLOGENIN ANALYSIS
Kapitel C.48 i dette bilag: Assayet for fisks reproduktion på kort sigt.
Care should be taken to avoid cross-contamination between VTG samples of males and females.
Se tillæg 4a.
Procedure 1A:
Findes her: [http://www.iso.org].
Fathead Minnow, Blood Collection from the Caudal Vein/Artery
Europa-Kommissionen (2007) Kommissionens henstilling 2007/526/EF af 18. juni 2007 om retningslinjer for anbringelse og pasning af dyr, der anvendes til test og andre videnskabelige formål (meddelt under nummer K(2007) 2525) ([http://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2007:197:0001:0089:DA:PDF]
After anaesthetisation, the caudal peduncle is partially severed with a scalpel blade and blood is collected from the caudal vein/artery with a heparinised microhematocrit capillary tube.
EU (2010) Europa-Parlamentets og Rådets direktiv 2010/63/EU af 22. september 2010 om beskyttelse af dyr, der anvendes til videnskabelige formål.
After the blood has been collected, the plasma is quickly isolated by centrifugation for 3 min at 15000 g (or alternatively for 10 min. at 15000 g at 4 °C).
Den EUT L 276 af 20.10.2010, s. 33-79.
If desired, percent haematocrit can be determined following centrifugation.
http://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2010:276:0033:0079:DA:PDF
The plasma portion is then removed from the microhematocrit tube and stored in a centrifuge tube with 0,13 units of aprotinin (a protease inhibitor) at – 80 °C until determination of VTG can be made.
Kapitel C.2 i dette bilag: Daphnia sp., Akut immobilisationstest.
Depending on the size of the fathead minnow (which is sex-dependent), collectable plasma volumes generally range from 5 to 60 microliters per fish (Jensen et al.
OECD, Paris.
2001).
Apikalt endepunkt
Procedure 1B:
Forårsager en virkning på populationsniveau.
Fathead Minnow, Blood Collection from Heart
En celledannelse rundt om dyrets poler, som omfatter en bestemt del af blommen.
Alternatively, blood may also be collected by cardiac puncture using a heparinised syringe (1000 units of heparin per ml).
Et stof eller en blanding
The blood is transferred into Eppendorf tubes (held on ice) and then centrifuged (5 min, 7000 g, room temperature).
Epiboli
The plasma should be transferred into clean Eppendorf tubes (in aliquots if the volume of plasma makes this feasible) and promptly frozen at – 80 °C, until analysed (Panter et al., 1998).
En massiv spredning af overvejende epidermale celler i embryonets gastrulationsfase og deres bevægelse fra den dorsale til den ventrale side, hvorved entodermale cellelag internaliseres i en invaginationslignende proces, og blommen indføres i embryonet.
Procedure 2A:
Gennemstrømningstest
Japanese Medaka, Excision of the Liver in Medaka
En test med en stadig strøm af testopløsninger gennem testsystemet under hele eksponeringen.
Removal of the test fish from the test chamber
Intern pladekontrol
Test fish should be removed from the test chamber using the small spoon-net.
Intern kontrol bestående af fire brønde fyldt med fortyndingsvand pr. plade med 24 brønde, som producenten eller forskeren bruger til at identificere potentiel forurening af pladerne under proceduren samt eventuel pladevirkning, som kan påvirke resultatet af testen (f.eks. temperaturgradient).
Be careful not to drop the test fish into other test chambers.
Vedligeholdelsesvand Det vand, som de voksne fisk dyrkes i.
In principle, the test fish should be removed in the following order: control, solvent control (where appropriate), lowest concentration, middle concentration, highest concentration and positive control.
Middel dødelig koncentration (LC50)
In addition, all males should be removed from one test chamber before the remaining females are removed.
Den koncentration af testkemikaliet, som skønnes at være dødelig for 50 % af testorganismerne i løbet af testens varighed.
The sex of each test fish is identified on the basis of external secondary sex characteristics (e.g. the shape of the anal fin).
Semistatisk udskiftningstest
Place the test fish in a container for transport and carry it to the workstation for excision of the liver.
Simplified Molecular Input Line Entry Specification.
If the sex cannot be identified by the fish's external appearance, remove all fish from the test chamber.
Somit
In this case, the sex should be identified by observing the gonad or secondary sex characteristics under a stereoscopic microscope.
I hvirveldyrembryoner under udvikling er somitter masser af mesoderm fordelt lateralt til neuralrøret, der i sidste ende vil udvikle dermis (dermatom), skeletmuskulatur (myotom) og ryghvirvler (sclerotom).
Excision of the liver
Statisk test
Transfer the test fish from the container for transport to the anaesthetic solution using the small spoon-net.
En test, hvor testopløsningerne ikke ændres i løbet af testen.
After the test fish is anesthetised, transfer the test fish on the filter paper (or a paper towel) using tweezers (commodity type).
Et stof eller en blanding, som testes ved hjælp af denne forsøgsmetode.
When grasping the test fish, apply the tweezers to the sides of the head to prevent breaking the tail.
Stoffer med ukendt eller variabel sammensætning, komplekse reaktionsprodukter eller biologiske materialer.
Wipe the water on the surface of the test fish on the filter paper (or the paper towel).
VEDLIGEHOLDELSE, AVL OG TYPISKE BETINGELSER FOR TEST AF AKUT TOKSICITET FOR ZEBRAFISKEMBRYONER
Then make a small transverse incision partway between the ventral neck region and the mid-abdominal region using dissection scissors.
Zebrafisk (Danio rerio)
Insert the dissection scissors into the small incision, and incise the abdomen from a point caudal to the branchial mantle to the cranial side of the anus along the midline of the abdomen.
Arternes oprindelse
Be careful not to insert the dissection scissors too deeply so as to avoid damaging the liver and gonad.
Indien, Burma, Malakka, Sumatra
Conduct the following operations under the stereoscopic microscope.
Kønsdimorfi
Place the test fish abdomen side up on the paper towel (glass Petri dish or slide glass are also available).
Hunner: fremspringende mave, når de bærer æg
Extend the walls of the abdominal cavity with precision tweezers and exteriorise the internal organs.
Hanner: mere slank, orange skær mellem blå langsgående striber (især synlige ved halefinnen)
It is also acceptable to exteriorise the internal organs by removing one side of the wall of the abdominal cavity if necessary.
Tørfoder (maks. 3 % af fiskens vægt pr. dag) 3-5 gange dagligt; derudover saltsøkrebs (Artemia spec.) nauplii og/eller små dafnier af passende størrelse fra en ikke-forurenet kilde.
Expose the connected portion of the liver and gallbladder using another pair of precision tweezers.
Fodring med levende foder er en kilde til miljøberigelse, og der bør derfor så vidt muligt anvendes levende foder.
Then grasp the bile duct and cut off the gallbladder.
Hanner: 0,5 ±0,1 g
Excise the caudal gastrointestinal tract from the anus and remove the tract from the abdominal cavity.
Vedligeholdelse af forældrefisk
Trim the mass of fat and other tissues from the periphery of the liver.
Lysstofrør (bredt spektrum); 10-20 μE/m2/s, 540-1080 lux eller 50-100 ft-c (laboratorieniveau); 12-16 timers lysperiode
Grasp the hepatic portal area using the precision tweezers and remove the liver from the abdominal cavity.
Yderligere vandkvalitetskriterier
Measure the liver weight with 1,5 ml microtube as a tare using an electronic analytical balance.
Partikler < 20 mg/l, total mængde organisk kulstof < 2 mg/l, total mængde organiske fosforpesticider < 50 ng/l, total mængde organiske chlorpesticider plus polychlorerede biphenyler < 50 ng/l
Record the value on the worksheet (read: 0,1 mg).
F.eks. 180 l, 1 fisk/l
Close the cap of the microtube containing the liver.
Fast (kulfiltreret); andre muligheder omfatter kombinationer med halvstatisk udskiftningsvedligeholdelse eller gennemstrømningssystem med kontinuerlig vandudskiftning
Following the excision of one liver, clean the dissection instruments or replace them with clean ones.
Anbefalede forhold mellem hanner og hunner til avl
Remove livers from all of the fish in the transport container as described above.
2:1 (eller massegydning)
After the livers have been excised from all of the fish in the transport container (i.e., all males or females in a test chamber), place all liver specimens in a tube rack with a label for identification and store it in a freezer.
Gydetanke
When the livers are donated for pre-treatment shortly after the excision, the specimens are carried to the next workstation in a cooling rack (or ice rack).
F.eks. 4-liters tanke forsynet med bund af stålgitter og planteattrap til at stimulere gydning; udvendige varmemåtter eller massegydning i vedligeholdelsestanke
Following liver excision, the fish carcass is available for gonad histology and measurement of secondary sex characteristics.
Æggenes struktur og udseende
Specimen
Stabil chorion (dvs. meget gennemsigtig, ikke-klæbrig, diameter ~ 0,8-1,5 mm)
Store the liver specimens taken from the test fish at ≤ – 70 °C if they are not used for the pre-treatment shortly after the excision.
Gydefrekvens
A cut is made just anterior to pectoral fins with scissors
En enkelt moden hunfisk gyder mindst 50-80 æg pr. dag.
The midline of abdomen is incised with scissors to a point approximately 2 mm cranial to the anus
Afhængigt af stammen kan gydefrekvensen være betydeligt højere.
The abdominal walls are spread with forceps for exposure of the liver and other internal organs
Befrugtningsfrekvensen bør være ≥ 70 %.
(Alternatively, the abdominal walls may be pinned laterally).
For fisk, der skal gyde første gang, kan befrugtningsfrekvensen for æggene være lavere i de første par gydninger.
Arrow shows liver
Statisk, semistatisk udskiftning, gennemstrømning, 26 ± 1 °C, 24 timers konditionerede testbeholdere (f.eks. plader med 24 brønde, 2,5-5 ml pr. brønd)
The liver is bluntly dissected and excised using forceps
NORMAL UDVIKLING HOS ZEBRAFISK VED 26 °C
Figure 5
Fig. 1: Udvalgte faser af tidlig udvikling hos zebrafisk (Danio rerio): 0,2-1,75 timer efter befrugtningen (fra Kimmel et al., 1995 (35)).
The intestines are gently retracted using forceps
Tidssekvensen for normal udvikling kan anvendes til at diagnosticere både befrugtning og æggenes levedygtighed (jf. punkt 26: Udvælgelse af befrugtede æg).
Figure 6
Fig. 2: Udvalgte faser af sen udvikling hos zebrafisk (Danio rerio) (embryoner uden chorion for optimal synlighed): 22-48 timer efter befrugtning (fra Kimmel et al., 1995 (35)).
Both ends of the intestines and any mesenteric attachments are severed using scissors
Fig. 3: Normal udvikling hos zebrafiskembryoner (Danio rerio): 1) 0,75 timer, 2-cellefase, 2) 1 time, 4-cellefase, 3) 1,2 timer, 8-cellefase, 4) 1,5 timer, 16-cellefase, 5) 4,7 timer, begyndende epiboli og 6) 5,3 timer, ca. 50 % epiboli (fra Braunbeck & Lammer 2006 (40)).
Figure 7 (female)
Layout af plader med 24 brønde
The procedure is identical for the female
1-5 fem testkoncentrationer/kemikalie;
Figure 8
nC negativ kontrol (fortyndingsvand),
The completed procedure
iC intern pladekontrol (fortyndingsvand)
Procedure 2 B:
pC positiv kontrol (3,4-DCA 4 mg/l);
Japanese Medaka (Oryzias latipes), Liver Pre-treatment for Vitellogenin Analysis
sC opløsningsmiddelkontrol
Take the bottle of homogenate buffer from the ELISA kit and cool it with crushed ice (temperature of the solution: ≤ 4°C).
æg pr. konc./ kontrol
If homogenate buffer from EnBio ELISA system is used, thaw the solution at room temperature, and then cool the bottle with crushed ice.
Gydningsenhed
Calculate the volume of homogenate buffer for the liver on the basis of its weight (add 50 μl of homogenate buffer per mg liver weight).
OVERSIGT OVER DØDELIGE ENDEPUNKTER FOR TESTEN FOR AKUT TOKSICITET FOR ZEBRAFISKEMBRYONER
For example, if the weight of the liver is 4,5 mg, the volume of homogenate buffer for the liver is 225 μl.
Følgende apikale endepunkter angiver akut toksicitet og dermed død for embryonerne: koagulering af embryonet, manglende løsnen af halen, manglende dannelse af somitter og manglende hjerteslag.
Prepare a list of the volume of homogenate buffer for all livers.
Følgende mikrografer er blevet udvalgt til at illustrere disse endepunkter.
Preparation of the liver for pre-treatment
Koagulering af embryonet
Take the 1,5 ml microtube containing the liver from the freezer just before the pre-treatment.
Under lysfeltbelysning viser koagulerede zebrafiskembryoner en række intransparente inklusioner.
Pre-treatment of the liver from males should be performed before females to prevent vitellogenin contamination.
Fig. 2
In addition, the pre-treatment for test groups should be conducted in the following order: control, solvent control (where appropriate), lowest concentration, middle concentration, highest concentration and positive control.
Manglende dannelse af somitter
The number of 1,5 ml microtubes containing liver samples taken from the freezer at a given time should not exceed the number that can be centrifuged at that time.
Selv om udviklingen forsinkes med ca. 10 timer, viser det 24 timer gamle zebrafiskembryon i a) veludviklede somitter (→), mens embryonet i b) ikke viser tegn på dannelse af somitter (→).
Arrange the 1,5 ml microtubes containing liver samples in the order of specimen number on the ice rack (no need to thaw the liver).
Selv om det 48 timer gamle zebrafiskembryon i c) klart viser et ødem i blommesækken (*), ses der tydeligt dannelse af somitter (→), mens det 96 timer gamle zebrafiskembryon i d) ikke udviser tegn på dannelse af somitter (→).
Operation of the pre-treatment
Bemærk også rygmarvskrumningen (scoliose) og det pericardiale ødem (*) vist i d).
Addition of the homogenate buffer
(a: →; 96 timer gammelt zebrafiskembryon).
Check the list for the volume of the homogenate buffer to be used for a particular sample of liver and adjust the micropipette (volume range: 100-1000 μl) to the appropriate volume.
Bemærk også det manglende øje (*).
Attach a clean tip to the micropipette.
Fig. 4
Take the homogenate buffer from the reagent bottle and add the buffer to the 1,5 ml microtube containing the liver.
Manglende hjerteslag er, pr. definition, vanskeligt at illustrere i en mikrograf.
Add the homogenate buffer to all of 1,5 ml microtubes containing the liver according to the procedure described above.
Manglende hjerteslag angives med manglende sammentrækninger i hjertet (dobbeltpil).
There is no need to change the micropipette tip to a new one.
Immobile blodceller i f.eks. aorta abdominalis (→ i indsats) er ikke indikator for manglende hjerteslag.
However, if the tip is contaminated or suspected to be contaminated, the tip should be changed.
Bemærk også den manglende dannelse af somitter i dette embryon (*, homogent snarere end segmentarisk udseende muskelvæv).
Homogenisation of the liver
Observationstiden for at registrere manglende hjerteslag bør være mindst et minut med en forstørrelse på mindst 80×.
Attach a new pestle for homogenisation to the microtube homogeniser.
C.50 Toksicitetstest af sedimentfri Myriophyllum spicatum
Insert the pestle into the 1,5 ml microtube.
Denne forsøgsmetode svarer til OECD-testvejledning (Test Guideline) 238 (2014).
Hold the microtube homogeniser to press the liver between the surface of the pestle and the inner wall of the 1,5 ml microtube.
Den er beregnet til at vurdere toksiciteten af kemikalier for Myriophyllum spicatum, en tokimbladet undervandsplante, som er en art i tusindbladsfamilien.
Operate the microtube homogeniser for 10 to 20 seconds.
Det er baseret på en eksisterende ASTM-forsøgsmetode (1), som er ændret som et sedimentfrit testsystem (2) for at anslå testkemikaliets iboende økotoksicitet (uafhængigt af testkemikaliets fordelingsadfærd mellem vand og sedimenter).
Cool the 1,5 ml microtube with crushed ice during the operation.