Med.Weter. strategies for the treatment of hypoxia-related ocular clonal antibody(Bs-1858r,Beijing,China)for one night at diseases (15,29).Despite all these results,there are 4C.The sections were rinsed in PBS and incubated at room no references for Ngb expression in the retina of the temperature for 80 minutes.The sections were re-washed Bactrian camel,rabbits,and sheep.The Bactrian camel and embedded in Trypsin(T-815025,Beijing,China).After (Bactrian camel)is a domestic animal that lives in that.the sections (70%alcohol (night).80%alcohol (5 h). desert and semi-desert areas and is highly adaptable 90%alcohol (3 h).95%alcohol (2 h).and n-butanol (night) to the extreme natural environment of drought and converted to n-butanol,(5 h)were prepared and embedded heat while adult rabbits are small mammals in the into paraffin wax.tissue sections were cut and examined family Leporidae and need a regulated environment to under an optical microscope (Olympus BX51). protect them against heat exhaustion or hypothermia. Reagents and instruments.The Trypsin(T-815025), Sheep are ruminant(cud-chewing)mammals of the Phosphate-buffered saline (PBS,Zli-9062)and rabbit genus Ovis and live throughout the world,especially anti-rat polyclonal antibody (Bs-1858r)were purchased in mountainous areas.The current research examined from Beijing Bio-Dee Biotechnology Co.Ltd.while the Ngb expression in the retina of healthy adult Bactrian immunohistochemical staining kit (sp-9001)and DAB color developing kit (Zli-9018)were ordered at Beijing camels,rabbits,and sheep by employing immuno- Zhongshan Jingiao Biotechnology Co.Ltd and Formalde- histochemical staining procedures.The research also hyde and n-butanol solution,analytically pure came from documented a theoretical foundation for understanding Tianjin Baishi Chemical Co.LTD.Glacial acetic acid and the repair function of Ngb in the retina injury of mam- alcohol solution:analysis-pure were bought from Tianjin mals under extreme natural environment and supplied Fuyu Fine Chemical Co Ltd and the normal sheep serum morphological data. (ab7489)came from North America.The following medical Material and methods instruments were purchased and employed during the sam- pling procedures:conventional surgical instruments which Animals and setting.The procedures used in this study include:scalpel and dissecting scissors.biological tissue were reviewed and approved by the Animal Ethics and Wel- embedding KD-BM machine,Bio-texturing and spread- fare Committee of Approval Number:Gansu Agricultural ing machine(K-P),and Biological tissue baking machine University (GAU-AEW-2019-0026). (KD-H)were purchased from Zhejiang Medical Getidy A total of three (3)healthy adult camels(7 years),five Medical Instruments Co.Zhejiang,Jinhua.The Constant (5)adult rabbits(6 years),and five(5)healthy adult sheep temperature incubator DHP-9082 was ordered at Shanghai (6 years)were involved in the study.The healthy adult Tajilin Industrial Co.Ltd,while the Semi-self-actuating camels were purchased from the Zhangye prefecture-level screw-cutting machine with a precision roller(Rm-2235) areas in Wuwei,Gansu province,China while the rabbits was bought from Leica,Germany,and Optical microscope and sheep used in this study were taken from Lanzhou (13395H2X)was purchased at Leica.Germany. city in Gansu Province.China were tested,and showed no Immuno-histochemical coloration.Streptavidin Per- sign of any disease.Specimens were collected in a morn- oxidase (SP)method was used for immunohistochemical ing and evening hour during July and collection was done staining while PBS was utilized as a negative control instead within one week.Animals were retrieved one after another of the primary antibody.Sheep serum was added to block from their housing area and taken to the slaughterhouse. the non-specific reactivity in each section and then observed Zhangye prefecture is located in central Gansu along the under the optical microscope (Olympus 13395H2X)to Hexi Corridor,occupying 42,000 km2(16,000 sq mi).It control color rendering.The color rendering period for takes up the entire breadth of the province,running from Inner Mongolia on the north to Qinghai on the south,but each section was consistent.After the reaction stopped by distilled water,the slices were re-dyed,dehydrated,and its urban core is at Ganzhou in the oasis formed by the Ruo transparent by hematoxylin and sealed.A Nikon Eclipse 80i, or Hei River and the Lanzhou city is the capital and larg- micro camera system was employed to capture specimen est city of Gansu Province and is noted for its yellow river images with consent shooting parameters and the Image- and situated in the temperate zone with a semi-arid climate during hot summer and extremely cold and dry winter.The Pro Plus 6.0 was introduced to acquire,count,measure,and average monthly temperature ranges from-5.3C(22.5F) classify optical density of each retina layer. Statistical analysis.Data analysis was performed using in January to 22.4C(72.3F)in July. Samples extraction and preparation.Before been Statistical Package Social Sciences(SPSS)1.0(1983)and 19.0(2009)to conduct one-way ANOVA,multiple com- sacrificed.all animals were fed and rested for a while.The animals were sacrificed,the cornea was open with a sharp parisons (Tamhane,ST2 method),and the Independent blade and the eyeballs were quickly removed and placed in Sample T-test was employed to evaluate the significant Davidson's fixation solution(95%alcohol,37%strong for- level.P<0.05 was considered significant.If there existed malin,glacial acetic acid,distilled water ratio:3:2:1:2) no significant difference in the MD,it was referred to as for 24 hours and stored at 4C.Later,the perfusion-fixed the negative control (P>0.05),and positive (+control or retinae were placed on disinfected glass slides,and normal expression if there was any significant difference. sheep serum(ab7489,North America)was added to block Results and discussion the non-specific binding sides at room temperature for an hour in Phosphate-buffered saline(PBS,Beijing,China). Adult Bactrian camel.The reports from HE stain- These sections were incubated with a rabbit anti-Ngb poly- ing showed that the Bactrian camel structure contains2 Med. Weter. strategies for the treatment of hypoxia-related ocular diseases (15, 29). Despite all these results, there are no references for Ngb expression in the retina of the Bactrian camel, rabbits, and sheep. The Bactrian camel (Bactrian camel) is a domestic animal that lives in desert and semi-desert areas and is highly adaptable to the extreme natural environment of drought and heat while adult rabbits are small mammals in the family Leporidae and need a regulated environment to protect them against heat exhaustion or hypothermia. Sheep are ruminant (cud-chewing) mammals of the genus Ovis and live throughout the world, especially in mountainous areas. The current research examined Ngb expression in the retina of healthy adult Bactrian camels, rabbits, and sheep by employing immuno￾histochemical staining procedures. The research also documented a theoretical foundation for understanding the repair function of Ngb in the retina injury of mam￾mals under extreme natural environment and supplied morphological data. Material and methods Animals and setting. The procedures used in this study were reviewed and approved by the Animal Ethics and Wel￾fare Committee of Approval Number: Gansu Agricultural University (GAU-AEW-2019-0026). A total of three (3) healthy adult camels (7 years), five (5) adult rabbits (6 years), and five (5) healthy adult sheep (6 years) were involved in the study. The healthy adult camels were purchased from the Zhangye prefecture-level areas in Wuwei, Gansu province, China while the rabbits and sheep used in this study were taken from Lanzhou city in Gansu Province, China were tested, and showed no sign of any disease. Specimens were collected in a morn￾ing and evening hour during July and collection was done within one week. Animals were retrieved one after another from their housing area and taken to the slaughterhouse. Zhangye prefecture is located in central Gansu along the Hexi Corridor, occupying 42,000 km2 (16,000 sq mi). It takes up the entire breadth of the province, running from Inner Mongolia on the north to Qinghai on the south, but its urban core is at Ganzhou in the oasis formed by the Ruo or Hei River and the Lanzhou city is the capital and larg￾est city of Gansu Province and is noted for its yellow river and situated in the temperate zone with a semi-arid climate during hot summer and extremely cold and dry winter. The average monthly temperature ranges from –5.3°C (22.5°F) in January to 22.4°C (72.3°F) in July. Samples extraction and preparation. Before been sacrificed, all animals were fed and rested for a while. The animals were sacrificed, the cornea was open with a sharp blade and the eyeballs were quickly removed and placed in Davidson’s fixation solution (95% alcohol, 37% strong for￾malin, glacial acetic acid, distilled water ratio: 3 : 2 : 1 : 2) for 24 hours and stored at 4°C. Later, the perfusion-fixed retinae were placed on disinfected glass slides, and normal sheep serum (ab7489, North America) was added to block the non-specific binding sides at room temperature for an hour in Phosphate-buffered saline (PBS, Beijing, China). These sections were incubated with a rabbit anti-Ngb poly￾clonal antibody (Bs-1858r, Beijing, China) for one night at 4°C. The sections were rinsed in PBS and incubated at room temperature for 80 minutes. The sections were re-washed and embedded in Trypsin (T-815025, Beijing, China). After that, the sections (70% alcohol (night), 80% alcohol (5 h), 90% alcohol (3 h), 95% alcohol (2 h), and n-butanol (night) converted to n-butanol, (5 h) were prepared and embedded into paraffin wax, tissue sections were cut and examined under an optical microscope (Olympus BX51). Reagents and instruments. The Trypsin (T-815025), Phosphate-buffered saline (PBS, Zli-9062) and rabbit anti-rat polyclonal antibody (Bs-1858r) were purchased from Beijing Bio-Dee Biotechnology Co. Ltd. while the immunohistochemical staining kit (sp-9001) and DAB color developing kit (Zli-9018) were ordered at Beijing Zhongshan Jinqiao Biotechnology Co. Ltd and Formalde￾hyde and n-butanol solution, analytically pure came from Tianjin Baishi Chemical Co. LTD. Glacial acetic acid and alcohol solution: analysis-pure were bought from Tianjin Fuyu Fine Chemical Co Ltd and the normal sheep serum (ab7489) came from North America. The following medical instruments were purchased and employed during the sam￾pling procedures: conventional surgical instruments which include: scalpel and dissecting scissors, biological tissue embedding KD-BM machine, Bio-texturing and spread￾ing machine (K-P), and Biological tissue baking machine (KD-H) were purchased from Zhejiang Medical Getidy Medical Instruments Co. Zhejiang, Jinhua. The Constant temperature incubator DHP-9082 was ordered at Shanghai Tajilin Industrial Co. Ltd, while the Semi-self-actuating screw-cutting machine with a precision roller (Rm-2235) was bought from Leica, Germany, and Optical microscope (13395H2X) was purchased at Leica, Germany. Immuno-histochemical coloration. Streptavidin Per￾oxidase (SP) method was used for immunohistochemical staining while PBS was utilized as a negative control instead of the primary antibody. Sheep serum was added to block the non-specific reactivity in each section and then observed under the optical microscope (Olympus 13395H2X) to control color rendering. The color rendering period for each section was consistent. After the reaction stopped by distilled water, the slices were re-dyed, dehydrated, and transparent by hematoxylin and sealed. A Nikon Eclipse 80i, micro camera system was employed to capture specimen images with consent shooting parameters and the Image￾Pro Plus 6.0 was introduced to acquire, count, measure, and classify optical density of each retina layer. Statistical analysis. Data analysis was performed using Statistical Package Social Sciences (SPSS) 1.0 (1983) and 19.0 (2009) to conduct one-way ANOVA, multiple com￾parisons (Tamhane, ST2 method), and the Independent Sample T-test was employed to evaluate the significant level. P < 0.05 was considered significant. If there existed no significant difference in the MD, it was referred to as the negative control (P > 0.05), and positive (+) control or expression if there was any significant difference. Results and discussion Adult Bactrian camel. The reports from HE stain￾ing showed that the Bactrian camel structure contains