Method for detecting protein loading capacity of porous calcium phosphate
A technology of porous calcium phosphate and detection method, which is applied in the detection field of protein load, can solve the problems of difficult determination of protein content, short-term release of protein, etc., achieve accurate detection, reduce the influence of hydrochloric acid, and reduce the influence of hydrochloric acid Effect
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Embodiment 1-4
[0049] Mix the rhBMP-2 porous calcium phosphate solidified product (abbreviation: calcium phosphate containing rhBMP-2) with 1M HCl (1mol / L HCl solution) in different proportions: 1g: 5mL (Example 1), 1g: 10mL (Example 2), 1g: 20mL (Example 3), and 1g: 30mL (Example 4) were dissolved, and it was observed within two minutes that the ratio of 1g: 5mL and 1g: 10mL contained rhBMP-2 Calcium phosphate cannot be completely dissolved, but at the ratio of 1g:20mL and 1g:30mL, calcium phosphate containing rhBMP-2 has been completely dissolved, the results are as follows figure 2 shown. figure 2 It is a diagram of the dissolution of calcium phosphate containing rhBMP-2 under different ratios according to the embodiments of the present application. Wherein A is the dissolution situation of embodiment 1, B is the dissolution situation of embodiment 2, C is the dissolution situation of embodiment 3, and D is the dissolution situation of embodiment 4.
[0050] When the dissolution ratio...
Embodiment 5
[0052] Weigh 100mg of BSA powder, add purified water, and dilute to 100mL to prepare a 1mg / mL BSA protein solution, which is diluted twice with water to obtain a 0.5mg / mL BSA protein solution. According to the ratio of 1g:20mL, add 2g of blank calcium phosphate to 40mL of 1M HCl, dissolve completely and set aside. Accurately measure 0, 10, 20, 30, 40, 50, and 60 μL of 0.5 mg / mL BSA protein solution with a pipette gun, add 1M HCl solution dissolved in blank calcium phosphate to make up to 400 μL, and configure a A series of protein concentration gradients (0, 12.5, 25, 37.5, 50, 62.5, 75 μg / mL), see Table 1 for specific operations.
[0053] Table 1 Standard protein solution preparation
[0054]
[0055] Grind the rhBMP-2-containing calcium phosphate to be tested into powder, dissolve it by adding 1M HCl at a ratio of 1g:20mL, pipette 40μL protein standard and each sample, add 200μL of Bradford working solution (accurate pipette measurement), set React in an incubator at 37...
Embodiment 6
[0061] Accurately weigh 200mg of BSA protein powder, add purified water, dilute to 10mL, mix well and set aside. According to the ratio of 1g:20mL, dissolve 0.2g of blank calcium phosphate in 4mL of 1M HCl, mix well and set aside.
[0062] Take 50 μL of 20 mg / mL BSA protein solution and add 950 μL of 1 M HCl solution in which blank calcium phosphate was dissolved to obtain a 1 mg / mL BSA protein concentration. Accurately measure 0, 20, 40, 60, 80, 100, 120 μL of 1 mg / mL BSA protein solution with a pipette gun, make up to 1000 μL with 1M HCl solution dissolved in blank calcium phosphate, mix well, and configure a series of protein Concentration gradient (0, 20, 40, 60, 80, 100, 120 μg / mL), see Table 3 for details.
[0063] Use a pipette to measure 30 μL standard BSA protein concentration gradient solution and each sample test solution (accurate), then add 300 μL Bradford working solution (accurate), react at room temperature for 5-10 minutes, and measure its concentration at 59...
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