natgillin/translations-raw huggingface dataset:
Potentially important characteristics of physical appearance in adult fathead minnows in endocrine disrupter testing include body colour (i.e. light/dark), coloration patterns (i.e. presence or absence of vertical bands), body shape (i.e. shape of head and pectoral region, distension of abdomen), and specialized secondary sex characteristics (i.e. number and size of nuptial tubercles, size of dorsal pad and ovipositor).
Če ločitev ni ustrezna, se suspenzija ponovno centrifugira v enakih pogojih.
Nuptial tubercles are located on the head (dorsal pad) of reproductively-active male fathead minnows, and are usually arranged in a bilaterally-symmetric pattern (Jensen et al.
Vsi vzorci se odstranijo iz ohlajene centrifuge in na rešetki za zamrzovanje razvrstijo po številki vzorca.
2001).
Paziti je treba, da se ločene plasti po centrifugiranju ne suspendirajo ponovno.
Control females and juvenile males and females exhibit no tubercle development (Jensen et al.
Odvzemanje supernatanta
2001).
Štiri 0,5-mililitrske mikroepruvete se za shranjevanje supernatanta položijo na rešetko za epruvete.
There can be up to eight individual tubercles around the eyes and between the nares of the males.
Z mikropipeto se odvzame 30 μl vsakega supernatanta (ločenega kot srednja plast) in prenese v eno 0,5-mililitrsko mikroepruveto.
The greatest numbers and largest tubercles are located in two parallel lines immediately below the nares and above the mouth.
Paziti je treba, da se ne odvzame lipid z vrhnje plasti ali jetrno tkivo s spodnje plasti.
In many fish there are groups of tubercles below the lower jaw;
Supernatant se odvzame in prenese v dve drugi 0,5-mililitrski mikroepruveti na enak način, kot je opisano zgoraj.
those closest to the mouth generally occur as a single pair, while the more ventral set can be comprised of up to four tubercles.
Z mikropipeto se odvzame preostali supernatant (če je mogoče: ≥ 100 μl).
The actual numbers of tubercles is seldom more than 30 (range, 18-28; Jensen et al.
Nato se supernatant prenese v preostali 0,5-mililitrski mikroepruveti.
2001).
Pokrov 0,5-mililitrske mikroepruvete se zapre in na nalepko zapiše količina supernatanta.
The predominant tubercles (in terms of numbers) are present as a single, relatively round structure, with the height approximately equivalent to the radius.
Takoj zatem se mikroepruvete ohladijo na rešetki za zamrzovanje.
Most reproductively-active males also have, at least some, tubercles which are enlarged and pronounced such that they are indistinguishable as individual structures.
Za vsak supernatant se konica mikropipete zamenja z novo.
Some types of endocrine-disrupting chemicals can cause the abnormal occurrence of certain secondary sex characteristics in the opposite sex;
Če se na konico prime veliko lipida, se takoj zamenja z novo, da se prepreči onesnaženje ekstrakta jeter z maščobo.
for example, androgen receptor agonists, such as 17β-methyltestosterone or 17β-trenbolone, can cause female fathead minnows to develop nuptial tubercles (Smith 1974; Ankley et al.
Ves centrifugirani supernatant se prenese v štiri 0,5-mililitrske mikroepruvete v skladu z zgoraj opisanim postopkom.
2001; 2003), while oestrogen receptor agonists may decrease number or size of nuptial tubercles in males (Miles-Richardson et al.
Ko se supernatant prenese v 0,5-mililitrske mikroepruvete, se te postavijo na rešetko za epruvete z identifikacijskimi nalepkami in takoj nato zamrznejo v zamrzovalniku.
1999; Harries et al.
Če se koncentracije VTG merijo takoj po predhodnem tretiranju, se ena 0,5-mililitrska mikroepruveta (ki vsebuje 30 μl supernatanta) ohrani hladna na rešetki za epruvete in prenese na delovno postajo, kjer se izvaja preskus ELISA.
2000).
V takem primeru se preostale mikroepruvete postavijo na rešetke za epruvete in zamrznejo v zamrzovalniku.
Below is a description of the characterization of nuptial tubercles in fathead minnows based on procedures used at the U.S. Environmental Protection Agency lab in Duluth, MN.
Po odvzemu supernatanta se ostanki primerno zavržejo.
Specific products and/or equipment can be substituted with comparable materials available.
Shranjevanje vzorca
Viewing is best accomplished using an illuminated magnifying glass or 3X illuminated dissection scope.
Dokler se 0,5-mililitrske mikroperuvete s supernatantom homogenata jeter ne uporabijo za ELISA, se shranijo pri ≤ – 70 °C.
View fish dorsally and anterior forward (head toward viewer).
Postopek 3A:
Place fish in small Petri dish (e.g., 100 mm in diameter), anterior forward, and ventral down.
Cebrica, odvzem krvi iz repne vene/arterije
Focus viewfinder to allow identification of tubercles.
Takoj po anesteziji se repno steblo prečno prereže, pri čemer se kri iz repne arterije/vene odvzame s heparinizirano mikrohematokritsko kapilarno cevko.
Gently and slowly roll fish from side to side to identify tubercle areas.
Količine krvi so v razponu od 5 do 15 μl in so odvisne od velikosti ribe.
Count and score tubercles.
V mikrokapilarno cevko se doda enaka količina pufra aprotinina (6 μgml v PBS), plazma pa se od krvi loči s centrifugiranjem (5 minut pri 600 g).
Repeat the observation on the ventral head surface by placing the fish dorsal anterior forward in the Petri dish.
Plazma se zbere v preskusnih epruvetah in shrani pri – 20 °C, dokler se ne analizira glede vitelogenina ali drugih zadevnih beljakovin.
Observations should be completed within 2 min for each fish.
Postopek 3B:
Tubercle Counting and Rating
Cebrica, odvzem krvi s srčno punkcijo
Six specific areas have been identified for assessment of tubercle presence and development in adult fathead minnows.
Da se prepreči koagulacija krvi in razgradnja beljakovin, se vzorci odvzamejo v fosfatnem pufru s soljo (PBS), ki vsebuje heparin (1000 enot/ml) in aprotinin, ki zavira proteazo (2 TIU/ml).
A template was developed to map the location and quantity of tubercles present (see end of this Appendix).
Za sestavine pufra se priporočajo heparin, amonijeva sol in liofilizirani aprotinin.
The number of tubercles is recorded and their size can be quantitatively ranked as:
Za vzorčenje krvi se priporoča brizga (1 ml) s pritrjeno tanko iglo (npr. Braun Omnikan-F).
0- absence, 1-present, 2-enlarged and 3-pronounced for each organism (Fig.
Brizga mora biti predhodno napolnjena s pufrom (približno 100 μl), da se iz vseh rib v celoti eluirajo majhne količine krvi.
1).
Vzorci krvi se odvzamejo s srčno punkcijo.
Rate 0- absence of any tubercle.
Najprej je treba ribe anestezirati z MS-222 (100 mg/l).
Rating 1-present, is identified as any tubercle having a single point whose height is nearly equivalent to its radius (diameter).
Ustrezna stopnja anestezije uporabniku omogoča, da loči srčni utrip cebric.
Rating 2- enlarged, is identified by tissue resembling an asterisk in appearance, usually having a large radial base with grooves or furrows emerging from the centre.
Med punkcijo srca naj bo bat brizge pod rahlo napetostjo.
Tubercle height is often more jagged but can be somewhat rounded at times.
Odvzeta količina krvi je v razponu med 20 in 40 mikrolitri.
Rating 3- pronounced, is usually quite large and rounded with less definition in structure.
Po srčni punkciji je treba zmes krvi/pufra prenesti v preskusno epruveto.
At times these tubercles will run together forming a single mass along an individual or combination of areas (B, C and D, described below).
Plazma se od krvi loči s centrifugiranjem (20 minut, 5000 g) in jo je treba shraniti pri – 80 °C, dokler se ne potrebuje za analizo.
Coloration and design are similar to rating 2 but at times are fairly indiscriminate.
cebrica, homogenizacija glave in repa
Using this rating system generally will result in overall tubercle scores of < 50 in a normal control male possessing a tubercle count of 18 to 20 (Jensen et al.
Ribe se anestezirajo in evtanazirajo v skladu z opisom preskusa.
2001).
Glava in rep ribe se odrežeta v skladu s sliko 1.
The actual number of tubercles in some fish may be greater than the template boxes (Appendix A) for a particular rating area.
Opozorilo:
If this happens, additional rating numbers may be marked within, to the right or to the left of the box.
med ravnanji s posameznimi ribami je treba vse instrumente za seciranje in rezalno desko sprati in ustrezno očistiti (npr. s 96-odstotnim etanolom), da se prepreči ‚vitelogeninsko onesnaženje‘ neinduciranih samcev zaradi samic ali induciranih samcev.
The template therefore does not need to display symmetry.
rez zadaj
An additional technique for mapping tubercles which are paired or joined vertically along the horizontal plane of the mouth could be done by double-marking two tubercle rating points in a single box.
rez za hrbtno plavutjo
Mapping regions:
za analizo VTG
A Tubercles located around eye.
za histologijo spol. žlez
Mapped dorsal to ventral around anterior rim of eye.
Teža zbranih glav in repov posameznih rib se izmeri na najbližji miligram.
Commonly multiple in mature control males, not present in control females, generally paired (one near each eye) or single in females exposed to androgens.
Po tehtanju se deli vnesejo v ustrezne epruvete (npr. 1,5-mililitrske epruvete) in zamrznejo pri – 80 °C do homogenizacije oz. dokler se neposredno ne homogenizirajo na ledu z dvema plastičnima pestiloma.
B Tubercles located between nares, (sensory canal pores).
epruvete je treba ustrezno oštevilčiti, da se lahko glava in rep ribe povežeta z ustreznim delom telesa, uporabljenim za histologijo spolnih žlez.
Normally in pairs for control males at more elevated levels (2- enlarged or 3- pronounced) of development.
Ko dobimo homogeno maso, se ji doda 4-kratna teža tkiva ledeno mrzlega homogenizacijskega pufra.
Not present in control females with some occurrence and development in females exposed to androgens.
Delo s pestiloma je treba nadaljevati, dokler zmes ni homogena.
C Tubercles located immediately anterior to nares, parallel to mouth.
Pomembno opozorilo:
Generally enlarged or pronounced in mature control males.
za vsako ribo se uporabita dve novi pestili.
Present or enlarged in less developed males or androgen-treated females.
Vzorci se postavijo na led do začetka centrifugiranja pri 4 °C in 50000 × g za 30 minut.
D Tubercles located parallel along mouth line.
S pipeto se supernatant odmeri na dele po 20 μl v vsaj dve epruveti, tako da se konica pipete potopi pod plast maščobe na površini in supernatant previdno izsesa brez delcev maščobe ali peletov.
Generally rated developed in control males.
Epruvete se do uporabe hranijo pri – 80 °C.
Absent in control females but present in androgen-exposed females.
Homogenizacijski pufer:(50 mM Tris-HCl pH 7,4; 1-odstotna zmes inhibitorja protaze (Sigma)):
E Tubercles located on lower jaw, close to mouth, usually small and commonly in pairs.
12 ml Tris-HCl pH 7,4 + 120 μl zmesi inhibitorja protaze.TRIS:
Varying in control or treated males, and treated females.
TRIS-ULTRA PURE (ICN), npr. iz Bie & Berntsen, Danska.Zmes inhibitorja protaze:
F Tubercles located ventral to E. Commonly small and paired.
Sigma (za tkivo sesalcev), številka izdelka P 8340.Opomba:
Present in control males and androgen-exposed females.
homogenizacijski pufer je treba uporabiti na dan priprave.
Tubercle Template
Med uporabo ga je treba postaviti na led.
Numerical Rating
Obogateni vzorci vitelogenina in medpreskusni referenčni standard
resent
Vsak dan izvedbe preskusov vitelogenina se analizira obogateni vzorec, pripravljen z medpreskusnim referenčnim standardom.
Date
Vitelogenin, uporabljen za pripravo medpreskusnega referenčnega standarda, je iz serije, ki se razlikuje od serije, uporabljene za pripravo kalibracijskih standardov za preskus, ki se izvaja.
nlarged
Obogateni vzorec se pripravi z dodajanjem znane količine medpreskusnega standarda vzorcu plazme kontrolnih samcev.
Total Score
Vzorec se obogati, da se doseže koncentracija vitelogenina, ki je med 10- in 100-krat večja od pričakovane koncentracije vitelogenina pri kontrolnih ribjih samcih.
ronounced
Vzorec plazme kontrolnih samcev, ki se obogati, lahko izvira iz ene ribe ali pa je sestavljen iz več rib.
Appendix 5B
Podvzorec neobogatene plazme kontrolnih samcev se analizira v vsaj dveh ponovitvenih vdolbinicah.
Assessment of secondary sex characteristics in medaka for the detection of certain endocrine active chemicals
Tudi obogateni vzorec se analizira v vsaj dveh ponovitvenih vdolbinicah.
Below is a description of the measurement of papillary processes, which are the secondary sex characteristics in medaka (Oryzias latipes).
Srednja količina vitelogenina v dveh neobogatenih vzorcih plazme kontrolnih samcev se doda izračunani količini vitelogenina, dodanega obogatenim vzorcem, da se določi pričakovana koncentracija.
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).
Razmerje med pričakovano koncentracijo in izmerjeno koncentracijo se sporoči skupaj z rezultati posameznih nizov preskusov, izvedenih isti dan.
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.
Tabela poteka za statistično analizo
3).
Dunnov ali Mann-Whitneyjev preskus s sredinami pon.
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.
statisTični diagram poteka za analizo razmerja med spoloma
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.
Transformacija za stabilizacijo?
When grasping the anal fin with tweezers, grasp a few fin rays in the anterior region with care not to scratch the papillary processes.
Dunn ali Mann-Whitneyjev preskus s sredinami pon.
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).
Transformacija za normalizacijo?
Measurement
Določite, ali je odziv na odmerek monoton.
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).
Transformacija za stabilizacijo varianc?
Place the fish abdomen side up.
Enake variance
Then cut the anal fin using small dissection scissors carefully (it is preferable to cut the anal fin with small amount of pterygiophore).
Nemonoton
Grasp the anterior region of the severed anal fin with tweezers and put it on a glass slide with a several drops of water.
Regresivni Jonckheerov ali Williamsov preskus
Then cover the anal fin with a cover glass.
Regresivni trendnostni preskus sredin ponovitev
Be careful not to scratch the papillary processes when grasping the anal fin with tweezers.
Enake variance Ne Da Da
Count the number of the joint plate with papillary processes using the counter under a biological microscope (upright microscope or inverted microscope).
Dunnov preskus s sredinami ponovitev
The papillary processes are recognized when a small formation of processes is visible on the posterior margin of joint plate.
Dunnov ali Mann-Whitneyjev preskus
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.
Dunnettov preskus ugnezdene analize varianc
If necessary, take a photograph of the anal fin and count the number of joint plate with papillary processes on the photograph.
Neenake variance
After the measurement, put the anal fin into the conical tube described in (1) and store it.
Sredine ponovitev so normalne in homogene.
Fig.1.
Opustite kontrolo z vodo
Diagram showing sexual difference in shape and size of the anal fin.
Regresivni Jonckheerov preskus
A, male;
Regresivni trendnostni preskus sredin ponovitev
B, female.
Ta preskusna metoda ustreza Smernici za preskušanje OECD (TG) 231 (2009).
Fig.2.A.
Preskus, s katerim se lahko zaznajo kemikalije, ki so aktivne v ščitničnem sistemu vretenčarjev, je treba razviti in validirati zaradi pomislekov, da bi lahko okoljske ravni kemikalij povzročile škodljive učinke za ljudi in prostoživeče živali.
Processes on joint plates of anal fin-ray.
OECD se je leta 1998 začela ukvarjati s prednostno nalogo revidiranja obstoječih smernic za preskušanje ter priprave novih smernic za presejalne preskuse in preskušanje potencialnih povzročiteljev endokrinih motenj.
J.P., joint plate;
Del naloge je bila priprava smernic za preskušanje za presejalno preskušanje kemikalij, ki so aktivne v ščitničnem sistemu vretenčarjev.
A.S., axial space;
Predlagana sta bila 28-dnevna študija oralne strupenosti s ponavljajočimi se odmerki na glodavcih (poglavje B.7 te priloge) in preskus preobrazbe dvoživk (AMA).
P., process.
Izboljšana preskusna metoda B.7 je prestala validacijo, objavljena pa je bila revidirana preskusna metoda.
B, Distal extremity of fin-ray.
Preskus preobrazbe dvoživk (AMA) je prestal validacijski program, ki je vključeval laboratorijske in medlaboratorijske študije, s katerimi sta bili dokazani ustreznost in zanesljivost preskusa (1) (2).
Actinotrichia (Act.) are on the tip.
Nato je validacijo preskusa strokovno pregledala skupina neodvisnih strokovnjakov (3).
Fig.3.
Ta preskusna metoda je rezultat izkušenj, pridobljenih med validacijskimi študijami za zaznavo kemikalij, ki vplivajo na ščitnico, in dela, opravljenega v državah članicah OECD.
Photograph of fish body showing the cut site when the gonad is fixed in the fixing solution other than 10 % neutral buffered formalin.
Preskus preobrazbe dvoživk (AMA) je presejalni preskus za empirično identifikacijo kemikalij, ki bi lahko vplivale na normalno funkcijo osi hipotalamus – hipofiza – ščitnica (HPT).
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.
AMA predstavlja posplošen model vretenčarja, saj temelji na ohranjenih strukturah in funkcijah osi HPT.
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).
Gre za pomemben preskus, ker je preobrazba dvoživk dobro proučen proces, odvisen od ščitnice, ki se odziva na kemikalije, aktivne v osi HPT, in je edini obstoječi preskus, s katerim se lahko zazna aktivnost ščitnice v živali, ki se morfološko razvija.
However, processes rarely appear on the first fin ray from the posterior end of the anal fin.
Splošni načrt poskusa zajema 21-dnevno izpostavljenost paglavcev Xenopus laevis v 51. fazi najmanj trem različnim koncentracijam preskusne kemikalije in kontroli z vodo za redčenje.
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).
Vsak preskusni tretirani vzorec ima štiri ponovitve.
Recommended procedures for sample collection for vitellogenin analysis
Gostota ličink na začetku preskusa je 20 paglavcev na preskusno posodo v vsaki tretirani skupini.
Care should be taken to avoid cross-contamination between VTG samples of males and females.
Opazovane končne točke so dolžina zadnjega kraka, dolžina od ust do zadnjične odprtine (SVL), razvojna faza, mokra teža, histologija ščitnice in dnevna opazovanja smrtnosti.
Procedure 1A:
Preskusna vrsta
Fathead Minnow, Blood Collection from the Caudal Vein/Artery
Xenopus laevis se rutinsko goji v laboratorijih po vsem svetu in se lahko enostavno pridobi prek komercialnih dobaviteljev.
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.
Razmnoževanje se lahko pri tej vrsti skozi celo leto enostavno doseže z uporabo injekcij humanega horionskega gonadotropina (HCG), pri čemer se lahko nastale ličinke v velikem številu rutinsko gojijo do izbranih razvojnih faz, zaradi česar je mogoča uporaba protokolov preskusa, specifičnih za razvojno fazo.
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).
Za uporabo v preskusu so primernejše ličinke odraslih osebkov iz laboratorija.
If desired, percent hematocrit can be determined following centrifugation.
Čeprav to ni prednostni postopek, se lahko namesto tega jajčeca ali embrii pošljejo v laboratorij, ki izvaja preskus, in aklimatizirajo;
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 vitellogenin can be made.
pošiljanje v stadiju ličink za uporabo v preskusu ni sprejemljivo.
Depending on the size of the fathead minnow (which is sex-dependent), collectable plasma volumes generally range from 5 to 60 microlitres per fish (Jensen et al.
Oprema in potrebščine
2001).
Za izvedbo tega preskusa so potrebne naslednja oprema in potrebščine:
Procedure 1B:
sistem za izpostavljanje (glej opis v nadaljevanju);
Fathead Minnow, Blood Collection from Heart
akvarij iz stekla ali nerjavnega jekla (glej opis v nadaljevanju);
Alternatively, blood may also be collected by cardiac puncture using a heparinized syringe (1000 units of heparin per ml).
posode za gojenje;
The blood is transferred into Eppendorf tubes (held on ice) and then centrifuged (5 min, 7000 g, room temperature).
naprave za nadzor temperature (npr. grelniki ali hladilniki (prilagodljivi do 22 °C ± 1 °C));
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 analyzed (Panter et al., 1998).
termometer;
Procedure 2A:
binokularni stereomikroskop;
Japanese Medaka, Excision of the Liver in Medaka
digitalni fotoaparat z ločljivostjo vsaj 4 milijone slikovnih točk in funkcijo mikro;
Removal of the test fish from the test chamber
programska oprema za digitalizacijo slik;
Test fish should be removed from the test chamber using the small spoon-net.
petrijevka (npr. 100 × 15 mm) ali prosojna plastična posodica primerljive velikosti;
Be careful not to drop the test fish into other test chambers.
analitska tehtnica, s katero je mogoče meriti na 3 decimalna mesta natančno (mg);
In principle, the test fish should be removed in the following order:
merilnik raztopljenega kisika;
control, solvent control (where appropriate), lowest concentration, middle concentration, highest concentration and positive control.
merilnik vrednosti pH;
In addition, all males should be removed from one test chamber before the remaining females are removed.
merilnik jakosti svetlobe, s katerim je mogoče meriti v luksih;
The sex of each test fish is identified on the basis of external secondary sex characteristics (e.g. the shape of the anal fin).
različna laboratorijska steklena posoda in pripomočki;
Place the test fish in a container for transport and carry it to the workstation for excision of the liver.
prilagodljive pipete (10 do 5000 μl) ali različne pipete ustreznih velikosti;
Check the labels of the test chamber and the transport container for accuracy and to confirm that the number of fish that have been removed from the test chamber and that the number of fish remaining in the test chamber are consistent with expectation.
preskusna kemikalija v zadostnih količinah za izvedbo študije, po možnosti iz ene serije;
If the sex cannot be identified by the fish's external appearance, remove all fish from the test chamber.
analitski instrumenti, primerni za preskušano kemikalijo, ali pogodbene analitske storitve.
In this case, the sex should be identified by observing the gonad or secondary sex characteristics under a stereoscopic microscope.
Kemijska preverljivost