Preparation method of FGF1 sericin protein gel with activity for promoting cell proliferation and product thereof
A technology of sericin and cell proliferation, applied in medical science, prostheses, etc., can solve the problems of weak cell adhesion, increased production cost, few products that have obtained clinical approval and finalization, and achieves strong water absorption performance and simple method Effect
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Embodiment 1
[0033] Example 1, the method for extracting sericin in human FGF1 silk
[0034] The cocoon shells were ground into powder by liquid nitrogen and used for later use; at a concentration of 50 mg / ml, extraction buffer 1 (50 mM Tris-HCl, 8 M urea, pH 7.0) was used for extraction at 80 ° C for 1 h; extraction buffer 2 ( 9.3M LiBr) at 60°C for 4h; using extraction buffer 3 (0.02M Na 2 CO 3 ) in a boiling water bath for 1 h to obtain a sericin solution containing human FGF1 protein.
Embodiment 2
[0035] Example 2, the preparation method of sericin gel (SS-FGF1) containing human FGF1 protein
[0036] Centrifuge the extract of extraction buffer 1 at 80°C, 13400rpm-18,000rpm for 30min, take the supernatant and refold the supernatant using the dithiothreitol / glutathione (DTT / GSSH) redox system FGF1 protein in . Refolding process: At 4°C, the supernatant was fully dialyzed with a dialysis bag (MWCO 1000Da, Spectrum Laboratory, Inc, USA) for 12 hours, and then replaced with a dialysate (8M urea, 1mM dithiothreitol (DTT), 50mM Tris–Cl ( pH 7.0), and 250mM NaCl); using half-dilution method using refolding solution (2.0mM reduced glutathione (GSH), 0.2mM oxidized glutathione (GSSG), 1mM DTT, 50mM Tris–Cl (pH 7.0), and 250mM NaCl ) to replace the dialysate, dialysis for 12 hours each time, repeat 4 times; dialyze with ultrapure water 6 times, remove the refolding solution, 12 hours each time. After renaturation is completed, the sericin aqueous solution is centrifuged at 4° C....
Embodiment 3
[0040] Example 3, performance detection of sericin gel containing human FGF1 protein
[0041] a. Electron microscope observation (SEM)
[0042] SS-FGF1 with a length and width of 1 cm was freeze-dried, and the inner layer was sprayed with gold, and observed and photographed with a scanning electron microscope (Supra 55sapphire, Zeiss). All experiments were performed at room temperature with a voltage of 3.0 kV. The number of samples in each experiment was 5 independent samples, and each sample was taken 3 times. SEM results showed that the interior of SS-FGF1 was a porous loose structure with a void size of 137±48 μm ( image 3 ).
[0043] b. Infrared spectroscopy (FTIR)
[0044] Freeze-dry the SS-FGF1 whose length and width are respectively 1 cm, and take its inner layer as a test sample. Determination was carried out using an infrared spectrometer (Thermo fisher scientific). An infrared spectrum analyzer (Thermo fisherscientific) was used to analyze the infrared absorp...
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