natgillin/translations-raw huggingface dataset:
Stock solutions of trace elements for medium M4 and M7
Zdroj variability v rámci laboratória a medzi laboratóriami je (najpravdepodobnejšie) založený na rôznych vývinových štádiách populácie rýb.
Stock solutions (I)
Vzhľadom na variabilitu merania VTG by sa k hodnotám NOEC založeným len na tomto sledovanom parametri malo pristupovať veľmi opatrne.
Amount (mg) made up to 1 litre of deionised water
Sú k dispozícii rôzne metódy na posúdenie tvorby VTG pri druhoch rýb použitých v tejto skúške.
To prepare the combined stock solution (II):
Pomerne citlivou a zároveň špecifickou meracou metódou je stanovenie koncentrácie bielkovín pomocou enzýmového imunosorbentového stanovenia (ELISA).
mix the following amounts (ml) of stock solutions (I) and make up to 1 litre of deionised water
Mali by sa použiť homologické protilátky (vytvorené proti VTG rovnakého druhu) a najdôležitejšie homologické štandardy.
Final concentrations in test solutions (mg/l)
Určenie pohlavia
MnCl2·4H2O [1]
V závislosti od postupu odberu vzoriek na analýzu VTG sa celá ryba alebo zvyšná stredná časť každej ryby umiestni do označenej spracovacej kazety a zafixuje vo vhodnom fixačnom prostriedku na účely histologického určenia pohlavia (voliteľne aj na vyhodnotenie štádia gonád).
SrCl2·6H2O [1]
Usmernenia na fixáciu a zaliatie sú uvedené v dodatku 7 ako aj v usmerňovacom dokumente OECD o diagnóze endokrinnej histopatológie gonád rýb (22).
MoO4·2H2O [1]
Po spracovaní sa ryby zalejú do parafínových blokov.
CuCl2·2H2O [1]
Ryby sa do parafínového bloku umiestňujú pozdĺžne.
CaCl2·6H2O
Z každého jedinca sa odoberie aspoň šesť pozdĺžnych rezov (s hrúbkou 3 – 5 μm) v čelnej rovine, ktoré obsahujú tkanivo z obidvoch gonád.
EDTA·2H2O [1] [2]
Rozstup medzi týmito rezmi by mal byť približne 50 μm pri samčekoch a 250 μm pri samičkách.
FeSO4·7H2O [1] [2]
Keďže však každý blok bude často obsahovať samčekov i samičky (ak je v každom bloku zaliatych viac jedincov), rozstup medzi rezmi z týchto blokov by mal byť približne 50 μm, kým sa nezíska aspoň šesť rezov gonád z každého samčeka.
Macro nutrient stock solutions for medium M4 and M7
Potom možno rozstup medzi rezmi zvýšiť na približne 250 μm pre samičky.
Amount made up to 1 litre of deionised water
Rezy sa zafarbia hematoxylínom a eozínom a vyhodnotia pomocou svetelnej mikroskopie s dôrazom na pohlavie (samčie, samičie, intersexuálne alebo nediferencované).
Amount of macro nutrient stock solutions added to prepare medium M4 and M7
Intersexuál je vymedzený ako prítomnosť viac, ako jedného oocytu v semenníkoch na šesť analyzovaných rezov alebo prítomnosť spermatogénnych buniek (áno/nie) vo vaječníkoch.
(ml/l)
Histopatologické vyšetrenie a stanovenie štádia vaječníkov a semenníkov je voliteľné, ale ak sa vykonáva, výsledky sa majú štatisticky analyzovať a uviesť do správy.
Final concentrations in test solutions M4 and M7
Treba poznamenať, že niektoré druhy rýb prirodzene nemajú plne vyvinutý pár gonád a môže byť prítomná len jedna gonáda (napr. medaka japonská a príležitostne dánio pruhované).
Vitamin stock solution for medium M4 and M7
Všetky tieto pozorovania sa musia zaznamenať.
All three vitamin solutions are combined to make a single vitamin stock solution.
Určenie genetického pohlavia pri jednotlivých medakách japonských je založené na prítomnosti alebo neprítomnosti génu určujúceho samčie pohlavie pri medakáchch DMY, ktorý sa nachádza na chromozóme Y.
Amount of vitamin stock solution added to prepare medium M4 and M7
Genotypové pohlavie medaky možno identifikovať sekvenovaním génu DMY z DNA vyextrahovanej napríklad z kúska análnej alebo chrbtovej plutvy.
Thiamine hydrochloride
Prítomnosť génu DMY znamená samčieho jedinca (XY) bez ohľadu na fenotyp, zatiaľ čo chýbajúci gén DMY znamená samičieho jedinca (XX) bez ohľadu na fenotyp (23).
Cyanocobalamin (B12)
Usmernenie na prípravu tkaniva a použitie metódy PCR sú uvedené v dodatku 9.
Biotine
Genetické pohlavie jednotlivých pichľaviek sivých sa takisto určuje metódou PCR, ktorá je opísaná v dodatku 10.
These substances differ in M4 and M7, as indicated above.
Výskyt intersexuálov (vymedzenie pojmu je uvedené v dodatku 1) je potrebné uviesť do správy.
These solutions are prepared individually, then poured together and autoclaved immediately.
Sekundárne pohlavné znaky
Preparation of formulated sediment
Pri druhoch ako medaka japonská sekundárne pohlavné znaky reguluje endokrinný systém.
SEDIMENT COMPOSITION
Pozorovania fyzického vzhľadu rýb by sa preto mali podľa možnosti uskutočniť na konci expozície.
The composition of the formulated sediment should be as follows:
V prípade medaky japonskej je bradavkový útvar v zadnej časti análnej plutvy samičiek citlivý na androgén.
Sphagnum moss peat, as close to pH 5,5-6,0 as possible, no visible plant remains, finely ground (particle size ≤ 1 mm) and air dried
Kapitola C.37 tejto prílohy (38) obsahuje príslušné fotografie samčích sekundárnych pohlavných znakov a androgenizovaných samičiek.
4 - 5
Je dôležité, aby sa sledovaný parameter vyhodnocoval pomocou najsilnejšieho platného štatistického testu.
> 50 % of the particles should be in the range of 50-200 μm
Pokusnou jednotkou je replikát, ale do štatistického testovania je potrebné zahrnúť aj variabilitu v rámci replikátu.
CaCO3, pulverised, chemically pure
V dodatku 8 je uvedená schéma rozhodovania ako pomôcka pri výbere najvhodnejšieho štatistického testu na základe povahy údajov získaných z testu.
0,05 - 0,1
Hladina štatistickej významnosti pre všetky zahrnuté sledované parametre je 0,05.
Conductivity ≤ 10 μS/cm
Podiel pohlaví a genetických pohlaví
PREPARATION
Podiely pohlaví sa majú analyzovať z hľadiska významného účinku (prístup NOEC/LOEC) expozície pomocou Jonckheereho-Terpstrovho (trendového) testu ak existuje monotónny priebeh krivky závislosti dávka-odpoveď.
The suspension is conditioned for at least two days with gentle stirring at 20 ± 2 °C, to stabilise pH and establish a stable microbial component. pH is measured again and should be 6,0 ± 0,5.
Ak sa zistí nemonotónnosť, je potrebné použiť párový test.
STORAGE
V ostatných prípadoch použite presný Mann-Whitneyho test s Bonferroniho-Holmovou úpravou.
The dry constituents for preparation of the artificial sediment may be stored in a dry and cool place at room temperature.
Dodatok 8 obsahuje schému opisujúcu štatistiku pre podiely pohlaví.
The formulated (wet) sediment should not be stored prior to its use in the test.
Podiely pohlaví sa musia uvádzať v tabuľkách ako podiely koncentrácií ± SD pre samčekov, samičky, intersexuálov a nediferencované jedince.
It should be used immediately after the 7 days conditioning period that ends its preparation.
Je potrebné uviesť štatistickú významnosť.
OECD (1984), Earthworm, Acute Toxicity Test, Test Guideline No. 207, Guidelines for the Testing of Chemicals, OECD, Paris.
Príklady sú uvedené vo validačnej správe o teste FSDT, fáza 2 (42).
Meller, M., Egeler, P., Roembke, J., Schallnass, H., Nagel, R. and B. Streit (1998), Short-term toxicity of lindane, hexachlorobenzene and copper sulfate on tubificid sludgeworms (Oligochaeta) in artificial media, Ecotox.
Genetické pohlavie sa uvádza ako percento zmeny fenotypového pohlavia pre samčekov, samičky, intersexuálov a nediferencované jedince.
Environ.
Koncentrácie VTG
Safety, 39:
Koncentrácie VTG sa majú analyzovať z hľadiska významného účinku (prístup NOEC/LOEC) expozície.
10-20.
Uprednostňuje sa Dunnettov test pred t-testom s Bonferroniho korekciou.
Chemical Characteristics of an Acceptable Dilution water
Ak sa použije Bonferroniho korekcia, uprednostňuje sa Bonferroniho-Holmova korekcia.
CONSTITUENT
Je možné urobiť logaritmickú transformáciu hodnôt VTG, aby sa dosiahla normalita a homogenita rozptylu.
CONCENTRATIONS
Ďalej ak je závislosť veličiny od koncentrácie v súlade s monotónnosťou, namiesto všetkých uvedených testov sa uprednostňuje Jonckheereho-Terpstrov test.
Hardness as CaCO3
Ak sa použijú t-testy alebo Dunnettov test, nie je potrebné dosiahnuť významnosť v F-teste analýzy ANOVA na to, aby bolo možné pokračovať.
However, it should be noted that if there is an interaction suspected between hardness ions and the test chemical, lower hardness water should be used (and thus, Elendt Medium M4 should not be used in this situation).
Podrobnosti nájdete v schéme v dodatku 8.
Guidance for test performance
Výsledky sa majú uvádzať v tabuľkách ako priemery koncentrácií ± SD samostatne pre samčekov, samičky, intersexuálov a nediferencované jedince.
Example of a breeding cage:
Je potrebné uviesť štatistickú významnosť pre fenotypové samičky a fenotypových samčekov.
A gauze on the top and at least one side of the cage (mesh size ca.
Skutočné koncentrácie testovanej chemikálie
1 mm)
Skutočné koncentrácie testovanej chemikálie v komorách sa majú analyzovať vo frekvenciách uvedených v bode 34.
B aperture for placing the emerged adults inside the breeding cage and to remove the laid egg ropes from the crystallisation dishes (not shown in this graphic)
Výsledky sa majú uvádzať v tabuľkách ako priemerná koncentrácia ± SD pre replikáty a pre jednotlivé koncentrácie, pričom treba uviesť informácie o počte vzoriek a extrémnych hodnotách odchyľujúcich sa od priemernej koncentrácie v jednotlivých skupinách o ± 20 %.
C breeding cage size minimum 30 cm length, 30 cm height and 30 cm width
Príklady sú uvedené vo validačnej správe o teste FSDT, fáza 2 (42).
Example of a test vessel:
Výsledky by sa mali interpretovať s opatrnosťou ak namerané koncentrácie testovanej chemikálie v testovacích roztokoch dosahujú úrovne blízke detekčnému limitu analytickej metódy.
A pasteur pipette for air supply of the overlying water
nominálne testované koncentrácie, priemery nameraných hodnôt a ich štandardné odchýlky v skúšobných komorách a metóda akou sa vypočítali (použitá analytická metóda by mala byť uvedená v prílohe), dôkaz o tom, že merania odkazujú na koncentrácie testovanej chemikálie v skutočnom roztoku,
B glass lid to prevent emerged midges from escaping
kvalita vody v skúšobných komorách:
C water surface layer
podrobné informácie o kŕmení (napr. druh potravy, zdroj, podávané množstvo a frekvencia a analýzy kontaminujúcich látok (napr. PCB, PAH a organochlórové pesticídy) ak to je relevantné.
D test vessel (glass beaker minimum 600 ml)
dôkaz, že kontrolné skupiny splnili kritériá validity:
E sediment layer
Údaje o prežití by sa mali uviesť ako percento na replikát a na koncentráciu.
Example of an exhauster for capturing adult midges (arrows indicate air flow direction):
Výsledky sa majú uviesť ako priemerné hodnoty ± štandardná odchýlka (SD) alebo štandardná chyba (SE).
A glass tube (inner diameter ca.
Štatistické údaje sa majú uvádzať minimálne ako hodnoty NOEC a LOEC a intervaly spoľahlivosti.
5 mm) connected to a self-priming pump B cork of vulcanised rubber, perforated with glass tube (A).
Je potrebné dodržiavať štatistickú schému (dodatok 8).
On the inside, the opening of glass tube (A) is covered with some cotton and a gauze (mesh size ca.
Sumpter, 1996, ‚Inhibition of testicular growth in rainbow trout (Oncorhynchus mykiss) exposed to estrogenic alkylphenolic chemicals‘, Environmental Toxicology and Chemistry 15, s.
1 mm) to prevent damaging the midges when they are sucked into the exhauster
194 – 202.
C transparent container (plastic or glass, length ca.
Sumpter, J.P. a S. Jobling, 1995, ‚Vitellogenesis As A Biomarker for Estrogenic Contamination of the Aquatic Environment‘, Environmental Health Perspectives 103, s.
15 cm) for captured midges
173 – 178.
D cork of vulcanised rubber, perforated with tube (E).
Hutchinson, S. Maddix a H. Trip (1999), ‚An in vivo testing system for endocrine disruptors in fish early life stages using induction of vitellogenin‘, Environmental Toxicology and Chemistry 18, s.
To release midges into the breeding cage, cork D is released from container C
337 – 347.
E tube (plastic or glass, inner diameter ca.
Pedersen a P. Bjerregaard (2001a), ‚Development of an ELISA for vitellogenin in whole body homogenate of zebrafish (Danio rerio)‘, Comparative Biochemistry and Physiology C-Toxicology & Pharmacology 130, s.
8 mm) to collect adult midges from vessel
119 – 131.
Schematic presentation of a life-cycle test:
Andersen, L., P. Bjerregaard a B. Korsgaard ( 2003), ‚Vitellogenin induction and brain aromatase activity in adult male and female zebrafish exposed to endocrine disrupters‘, Fish Physiology and Biochemistry 28, s.
A 1st generation — test vessels containing a sediment-water system, eight replicates, 20 first instar larvae per vessel
319 – 321.
B four test vessels for each breeding cage, A and B
Petersen (2003), ‚Gonad development and vitellogenin production in zebrafish (Danio rerio) exposed to ethinylestradiol and methyltestosterone‘ aquatic Toxicology 65, s.
C breeding cages (A and B) for swarming, mating and oviposition
397 – 411.
D crystallising dishes for deposition of egg ropes
Tyler (2002), ‚Utility of a juvenile fathead minnow screening assay for detecting (anti-)estrogenic substances‘, Environmental Toxicology and Chemistry 21, s.
E micro plates, one well for each egg rope
319 – 326.
F 2nd generation — test vessels containing a sediment-water system, eight replicates, 20 first instar larvae per vessel
Wang (2007), ‚Toxicity of the aromatase inhibitor letrozole to Japanese medaka (Oryzias latipes) eggs, larvae and breeding adults‘, Comparative Biochemistry and Physiology C-Toxicology & Pharmacology 145, s.
C.41.
533 – 541.
FISH SEXUAL DEVELOPMENT TEST
LeBlanc a C.V.Sullivan (1999), ‚Fathead minnow (Pimephales promelas) vitellogenin: purification, characterization and quantitative immunoassay for the detection of estrogenic compounds‘, Comparative Biochemistry and Physiology C-Toxicology & Pharmacology 123, s.
This test method is equivalent to OECD test guideline (TG) 234 (2011).
113 – 125.
It is based on a decision from 1998 to develop new or update existing test methods for the screening and testing of potential endocrine disrupters.
Porcher (2002), ‚Development and validation of an enzyme-linked immunosorbent assay to measure vitellogenin in the zebrafish (Danio rerio)‘, Environmental Toxicology and Chemistry 21, s.
The Fish Sexual Development Test (FSDT) was identified as a promising test method covering a sensitive fish life stage responsive to both oestrogen and androgen-like chemicals.
1699 – 1708.
The test method went through an inter-laboratory validation exercise from 2006 to 2010, where Japanese medaka (Oryzias latipes), zebrafish (Danio rerio) and three spined stickleback (Gasterosteus aculeatus) were validated and fathead minnow (Pimephales promelas) was partially validated (41) (42) (43).
Hatano, H. Mizukami, M. Yamashita, R. Sakakibara a E. Tamiya (2002), ‚Development and application of a monoclonal antibody-based sandwich ELISA for quantification of Japanese medaka (Oryzias latipes) vitellogenin‘, Comparative Biochemistry and Physiology C-Toxicology & Pharmacology 132, s.
This protocol includes Japanese medaka, the three-spined stickleback and zebrafish.
161 – 169.
The protocol is in principle an enhancement of OECD TG 210 Fish, Early Life Stage Toxicity Test (1), where the exposure is continued until the fish are sexually differentiated, i.e. about 60 days post-hatch (dph) for Japanese medaka, the three-spined stickleback and zebrafish (the exposure period can be shorter or longer for other species that are validated in the future), and endocrine-sensitive endpoints are added.
Bengtsson (2004), ‚The juvenile three-spined stickleback (Gasterosteus aculeatus L.) as a model organism for endocrine disruption – II – kidney hypertrophy, vitellogenin and spiggin induction‘ aquatic Toxicology 70, s.
The combination of the two core endocrine endpoints, vitellogenin (VTG) concentration and phenotypic sex ratio enable the test to indicate the mode of action of the test chemical.
Tatarazako, N., M. Koshio, H. Hori, M. Morita a T. Iguchi (2004), ‚Validation of an enzyme-linked immunosorbent assay method for vitellogenin in the medaka‘, Journal of Health Science 50, s.
Due to the population-relevant change in phenotypic sex ratio, the FSDT can be used for hazard and risk assessment.
301 – 308.
However, if the test is used for hazard or risk assessment, the stickleback should not be used because the validation data available so far showed that in this species the alterations of phenotypic sex ratio by the test chemicals were uncommon.
Eidem, J.K., H. Kleivdal, K. Kroll, N. Denslow, R. van Aerle, C. Tyler, G. Panter, T. Hutchinson a A. Goksoyr (2006), ‚Development and validation of a direct homologous quantitative sandwich ELISA for fathead minnow (Pimephales promelas) vitellogenin.
The protocol is based on fish exposed via water to chemicals during the sex labile period in which the fish is expected to be most sensitive to the effects of endocrine disrupting chemicals that interfere with sexual development.
Aquatic Toxicology‘, 78, s.
Two core endpoints are measured as indicators of endocrine-associated developmental aberrations, the VTG concentrations and sex ratios (proportions of sex) determined via gonad histology.
202 – 206.
Gonadal histopathology (evaluation and staging of oocytes and spermatogenetic cells) is optional.
Ankley (2006), ‚Evaluation of a commercial kit for measuring vitellogenin in the fathead minnow (Pimephales promelas)‘, Ecotoxicology and Environmental Safety 64, s.
Additionally, the genetic sex is determined whenever possible (e.g. in Japanese medaka and the three spined stickleback).
101 – 105.
The presence of a genetic sex marker is a considerable advantage as it increases the power of the sex ratio statistics and enables the detection of individual phenotypic sex reversal.
Holbech, H., Petersen, G. I., Norman a., Örn, S, Norrgren, L. a Bjerregaard, P (2001b), ‚Suitability of zebrafish as test organism for detection of endocrine disrupting chemicals.
Other apical endpoints that should be measured include hatching rate, survival, length and body weight.
Comparison of vitellogenin in plasma and whole body homogenate from zebrafish (Danio rerio) and rainbow trout (Oncorhynchus mykiss)‘, Nordic Council of Ministers, TemaNord 2001:597, s.
The test method might be adaptable to other species than those mentioned above provided that the other species undergo a validation equal to the one accomplished for Japanese medaka, the three-spined stickleback and zebrafish, that the control fish are sexually differentiated at the end of the test, that VTG levels are sufficiently high to detect significant chemical-related variations, and that sensitivity of the test system is established using endocrine active reference chemicals ((anti)-oestrogens, (anti)-androgens, aromatase inhibitors etc).
48 – 51.
In addition, any validation report(s) referring to FSDT data using other species should be reviewed by the OECD, and the validation outcome should be considered as satisfactory.
Porcher a A. Goksoyr (2004), ‚Development of quantitative vitellogenin-ELISAs for fish test species used in endocrine disruptor screening‘ analytical and Bioanalytical Chemistry 378, s.
Initial considerations and limitations
621 – 633.
VTG is normally produced by the liver of female oviparous vertebrates in response to circulating endogenous oestrogen (2).
Orn, S., S. Yamani a L. Norrgren (2006), ‚Comparison of vitellogenin induction, sex ratio and gonad morphology between zebrafish and Japanese medaka after exposure to 17 alpha-ethinylestradiol and 17 beta-trenbolone‘ archives of Environmental Contamination and Toxicology 51, s.
The VTG synthesis is very limited, though detectable, in immature fish and adult male fish because they lack sufficient circulating oestrogen.
237 – 243.
However, the liver is capable of synthesising and secreting VTG in response to exogenous oestrogen stimulation (3) (4) (5).
Scholz, S. a N. Kluver (2009), ‚Effects of Endocrine Disrupters on Sexual, Gonadal Development in Fish‘, Sexual Development 3, s.
The measurement of VTG serves for the detection of chemicals with oestrogenic, anti-oestrogenic, androgenic modes of action and chemicals that interfere with steroidogenesis as for example aromatase inhibitors.
136 – 151.
The detection of oestrogenic chemicals is possible via the measurement of VTG induction in male fish, and it has been abundantly documented in the scientific peer-reviewed literature.
Fenske, M., G. Maack, C. Schafers a H. Segner (2005), ‚An environmentally relevant concentration of estrogen induces arrest of male gonad development in zebrafish, Danio rerio‘, Environmental Toxicology and Chemistry 24, s.
VTG induction has also been demonstrated following exposure to aromatisable androgens (6) (7).
1088 – 1098.
A reduction in the circulating level of oestrogen in females, for instance through the inhibition of the aromatase converting the endogenous androgen to the natural oestrogen 17β-oestradiol, causes a decrease in the VTG concentration, which is used to detect chemicals having aromatase inhibiting properties or steroidogenesis inhibitors more broadly (33).
Nagahama (2004), ‚Two DM domain genes, DMY and DMRT1, involved in testicular differentiation and development in the medaka, Oryzias latipes‘, Developmental Dynamics 231, s.
The biological relevance of the VTG response following oestrogenic/aromatase inhibition is established and has been broadly documented (8) (9).
518 – 526.
However, it is possible that production of VTG in females can also be affected by general toxicity and non-endocrine toxic modes of action.
Shinomiya a., H. Otake, K. Togashi, S. Hamaguchi a M. Sakaizumi (2004), ‚Field survey of sex-reversals in the medaka, Oryzias latipes: genotypic sexing of wild populations‘, Zoological Science 21, s.
Several measurement methods have been successfully developed and standardised for routine use to quantify VTG in blood, liver, whole body or head/tail homogenate samples collected from individual fish.
613 – 619.
This is the case for zebrafish, three-spined stickleback and Japanese medaka and also the partially validated species fathead minnow;
Flick (2007), ‚Collapse of a fish population after exposure to a synthetic estrogen‘, Proceedings of the National Academy of Sciences of the United States of America 104, s.
species-specific Enzyme-Linked Immunosorbent Assay (ELISA) methods using immunochemistry for the quantification of VTG are available (5) (10) (11) (12) (13) (14) (15) (16).
8897 – 8901.
In Japanese medaka and zebrafish, there is a good correlation between VTG measured from blood plasma, liver and homogenate samples although homogenates tend to show slightly lower values than plasma (17) (18) (19).
Kidd (2009) ‚Interspecies differences in biochemical, histopathological, and population responses in four wild fish species exposed to ethynylestradiol added to a whole lake‘, Canadian Journal of Fisheries and Aquatic Sciences 66, s.
Appendix 5 provides the recommended procedures for sample collection for VTG analysis.
1920 – 1935.
Change in the phenotypic sex ratio (proportions of sex) is an endpoint reflecting sex reversal.
Tyler (2006), ‚Development of chronic tests for endocrine active chemicals – Part 1.
In principle, oestrogens, anti-oestrogens, androgens, anti-androgens and steroidogenesis inhibiting chemicals can affect the sex ratio of developing fish (20).
An extended fish early-life stage test for oestrogenic active chemicals in the fathead minnow (Pimephales promelas)‘, Aquatic Toxicology 77, s.
It has been shown that this sex reversal is partly reversible in zebrafish (21) following oestrogen-like chemical exposure, whereas sex reversal following androgen-like chemical exposure is permanent (30).
279 – 290.
The sex is defined as female, male, intersex (both oocytes and spermatogenetic cells in one gonad) or undifferentiated, determined in individual fish via histological examination of the gonads.
Petersen, P. Jackson, K. Hylland, L. Norrgren a P. Bjerregaard (2006), ‚Detection of endocrine disrupters: Evaluation of a Fish Sexual Development Test (FSDT)‘, Comparative Biochemistry and Physiology C-Toxicology & Pharmacology 144, s.
Guidance is given in Appendix 7 and in the OECD Guidance Document on the Diagnosis of Endocrine-Related Histopathology of Fish Gonads (22).
57 – 66.