Leech fibrinolytic protein, preparation method and application thereof
A technology of fibrinolytic protein and leech, which is applied in the field of medicine and achieves the effect of great development and application prospects
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0019] Embodiment 1 extracts leech fibrinolytic protein from dry leech
[0020] Proceed as follows:
[0021] (1) Weigh the dried leeches, pulverize them with a pulverizer, add 20 times of water and soak them for 48 hours at 4°C and then homogenize them. Add 1% trypsin and 5% crude enzyme of dry leeches to the homogenate for enzymolysis (pH8.2~8.5, temperature 50°C), after 5h, trypsin with 1% weight of dry leech was added to continue enzymolysis for 4h; after enzymolysis was completed, the temperature returned to room temperature, and absolute ethanol was added to the enzymolysis solution to 75% , left standing overnight (12 hours) at 4°C, suction filtered to obtain the supernatant, concentrated under reduced pressure (concentrated to one-tenth of the original volume) to obtain the crude leech protein extract;
[0022] (2) The above-mentioned leech fibrinolytic protein crude product extract is carried out QSephroseFastFlow ion-exchange column chromatography, with the NaCl solu...
Embodiment 2
[0024] Example 2 The dissolution of leech fibrinolytic protein to fibrin
[0025] Specific steps are as follows:
[0026] Preparation of fibrin plate: Weigh 125mg of agarose, add it to 23ml of normal saline, boil and swell and balance it in a water bath at 55°C to 60°C; add 14ul100U / ml thrombin and 2.2ml6mg / ml fibrinogen solution after balance , shake continuously until cloudy, pour it into a plate with a diameter of 8cm immediately after being cloudy, place it horizontally and fully solidify, place it at 4°C for 30min, and then punch holes.
[0027] Determination of fibrinolytic activity: add 10ul leech fibrinolytic protein solution (the leech fibrinolytic protein prepared in embodiment 1, the concentration of leech fibrinolytic protein solution is 0.87mg / ml) in the well, after 37 ℃ incubation 24h, to fibrin The dissolution situation was observed and measured. Fibrinolysis results such as image 3 shown by image 3 It can be seen that the crude extract of leech fibrinolyt...
Embodiment 3
[0028] Example 3 The effect of leech fibrinolytic protein on thrombus dissolution in model mice
[0029] Specific steps are as follows:
[0030] 40 Kunming mice (body weight 20±1.0 g) were randomly divided into 4 groups, 10 mice in each group, half male and half female, and cultured separately. The blank control group, the model control group, the leech fibrinolytic protein crude extract group, and the HAP-5-1 group were followed in turn.
[0031] Except for the blank control group, thrombus models were produced in the other three groups. Slightly anesthetize the mice with ether, sterilize the back with 70% ethanol, inject 10 mg / mL carrageenan solution subcutaneously in the back at a dose of 100 mg / kg, and place the mice in a room with a temperature lower than 18°C after the injection is completed. Feed in the environment, and pay attention to the changes in the tail of the mice. A dark red thrombosis area appeared on the tip of the tail of the mice during 4-24 hours, and...
PUM
| Property | Measurement | Unit |
|---|---|---|
| Molecular weight | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
Login to View More