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Method for building oxidative stress model of carp red blood cells

A technology of oxidative stress and model building, applied in the biological field, can solve problems such as unstable concentration and poor controllability of modeling effect

Inactive Publication Date: 2014-04-16
SICHUAN AGRI UNIV
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

But H 2 o 2 In the cell culture medium, the metabolism is fast, the concentration is unstable, and the controllability of the modeling effect is poor.

Method used

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  • Method for building oxidative stress model of carp red blood cells
  • Method for building oxidative stress model of carp red blood cells
  • Method for building oxidative stress model of carp red blood cells

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Embodiment 1

[0027] Embodiment 1: A method for establishing a fish erythrocyte oxidative stress model, the following steps are performed in sequence:

[0028] 1. Disinfect 3 carps weighing about 150 grams according to conventional methods, and draw about 1 ml of blood from the tail of each fish with a syringe.

[0029] 2. Put the blood into 6 ml carp normal saline containing 40 IU / ml sodium heparin, centrifuge the blood sample at 900 g, 4° C. for 10 minutes, and discard the supernatant.

[0030] 3. The erythrocyte pellet was suspended in 6 ml carp normal saline, and then centrifuged at 900 g, 4°C for 10 minutes.

[0031] 4. Resuspend the red blood cell pellet with carp saline, wash twice according to step 3, suspend the cell pellet in 6 ml of carp saline, and store at 4°C for later use.

[0032] 5. Take an appropriate amount of red blood cell suspension and add an equal volume of Fe 2+ / H 2 o 2 Mix the solutions with final concentrations of 0 / 0, 10 μM / 5 μM, 20 μM / 10 μM, 30 μM / 15 μM, 40...

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Abstract

The invention discloses a method for building an oxidative stress model of carp red blood cells, which comprises sterilizing the tail of a carp, drawing about 1 millimeter of blood from the tail of each carp; putting the blood into 6 millimeters of carp saline solution containing heparin, centrifuging 900 grams of a blood sample at a temperature of 4 DEG C for 10 minutes, rejecting supernate; suspending red blood cell deposits in the 6 millimeters of carp saline solution, centrifuging 900 grams of sample at a temperature of 4 DEG C for 10 minutes; re-suspending the red blood cell deposits in the carp saline solution, washing suspending the red blood cell deposits in the 6 millimeters of carp saline solution after the red blood cell deposits are washed twice, preserving the red blood cell deposits at a temperature of 4 DEG C for reserve; drawing adequate red blood cell suspension, adding equal volume of Fe2+ mixture solution with a density of 40 mu M or H2O2 mixture solution with a density of 20 mu M, incubating the suspension at a temperature of 37 DEG C for 8 to 10 hours; centrifuging 1000 grams of sample at a temperature of 4 DEG C for 3 minutes, and detecting apoptosis of cell samples by a flow cytometry and agarose gel electrophoresis. The method for building the oxidative stress model of carp red blood cells utilizes the Fe2+ or H2O2 mixture solution as a source of free radical OH, for the first time, and a red blood cell oxidative stress model is built.

Description

technical field [0001] The invention relates to a method for establishing a fish erythrocyte oxidative stress model, which belongs to the field of biotechnology. Background technique [0002] Oxidative damage caused by free radicals (singlet oxygen, superoxide anion, hydroxyl radicals) is the cause of decline in fish health and various diseases. At the same time, free radicals can directly damage the biological macromolecules of muscle tissue, such as nucleic acid, protein, and lipid, and reduce the quality of fish products. Therefore, it is very important to study the oxidative damage and anti-oxidation mechanism of fish and improve the anti-oxidation capacity of fish for the healthy breeding of fish and the quality of fish products. [0003] Red blood cells have important functions such as maintaining internal environment stability, transportation, regulation, and defense. But the environment in which red blood cells live is constantly exposed to free radicals. Red bloo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/078
Inventor 周小秋李华涛冯琳刘扬姜俊姜维丹
Owner SICHUAN AGRI UNIV
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