A Method for Evaluating Components Alleviating Foodborne Enteritis Based on a Juvenile Zebrafish Imaging Model
An imaging model, zebrafish technology, applied in fish farming, analytical materials, food processing, etc., can solve the problems of large differences and weaken the response of intestinal immune cells, so as to shorten the evaluation period, reliable evaluation methods, and overcome analysis step complex effect
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Embodiment 1
[0035] Example 1: Application of Imaging of Innate Immunofluorescence-labeled Juvenile Zebrafish Strains in Analyzing the Effect of Sinomenine on Alleviating Soybean Meal-Induced Enteritis
[0036] (1) Preparation of double transgenic zebrafish Tg(lyz:DsRED2 / mpeg1:EGFP)
[0037]1) Feed the transgenic zebrafish Tg(lyz:DsRED2) and Tg(mpeg1:EGFP) that have been reared separately one week in advance, and feed them three times a day for half an hour. The temperature of the breeding water is controlled at 28±0.5°C, and the light time is 14h , dark for 10 hours, the environment of the zebrafish breeding system is pH: 7.0-8.0, salinity 0.25-0.50‰, dissolved oxygen 5-8mg / L, total ammonia nitrogen less than 0.02mg / L, other conditions refer to the National Zebrafish Breeding Center Request (http: / / www.zfish.cn);
[0038] 2) One female and one male corresponding to the fluorescent strains of Tg(lyz:DsRED2) and Tg(mpeg1:EGFP) were paired overnight in a matching fish tank, and the separato...
Embodiment 2
[0071] Example 2: Application of imaging of juvenile zebrafish strains with fluorescently labeled lymphocytes in analyzing the effect of sinomenine on alleviating SBMIE
[0072] (1) Maintenance of fish mating and film-forming seedlings
[0073] 1) Preparation before fish mating: Feed the transgenic zebrafish Tg(rag2:DsRed) and Tg(lck:lck-eGFP) one week in advance, and feed them three times a day for half an hour. The time is 14h, dark 10h, all the other cultivation conditions and experimental steps are the same as (1) in embodiment 1; rag2 marks the immature (divide) lymphocyte, lck is the mark of T lymphocyte maturation, and the main lymphocyte in the intestine taxa;
[0074] 2) Test cross to obtain fluorescent experimental fish: cross Tg(rag2:DsRed) and Tg(lck:lck-eGFP) homozygous adult fish with wild-type zebrafish, one female and one male were paired overnight in a mating tank, and the second Unplug the separator every morning, let it spawn naturally for one hour, collec...
Embodiment 3
[0088] Example 3: Application of imaging of larval zebrafish intestinal slices in analyzing the effect of sinomenine on alleviating SBMIE
[0089] (1) Sampling and fixing, dehydration and wax immersion treatment
[0090] 1) Get the zebrafish fry after 27pdf of corresponding different groups of treatments (the fry of the acquired immune model is shown in Example 2), gently put the fry into 4% paraformaldehyde fixative solution with tweezers, and take two for each group. Paraffin section of the hindgut was carried out;
[0091] 2) Take out the whole fish, smooth it and put it into the dehydration box, put the dehydration box into the hanging basket and dehydrate it in the dehydrator;
[0092] 3) Dehydration: 75% alcohol 4h-85% alcohol 2h-90% alcohol 2h-95% alcohol 1h-absolute ethanol I 30min-absolute ethanol II 30min;
[0093] 4) Transparent: put ethanol after dehydration: toluene (1:1) 5-10min-xylene I 5-10min-xylene II 5-10min;
[0094] 5) Through the wax: 65 ℃ melting para...
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