Serum-free culture medium for stem cells
A technology of serum-free medium and basal medium, applied in the biological field, can solve the problems of insufficient types of Rho kinase inhibitors and antibody inhibitors, etc., to achieve inhibition of invasion and migration, good application value, and promotion of proliferation Effect
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Embodiment 1
[0031] Example 1 Preparation of Rho GTPase monoclonal antibody
[0032] Rho GTPase 1 (RND1) was used as the immunogen (Shanghai Hewu Biotechnology Co., Ltd. purchased in May 2020).
[0033] Using Rho GTPase 1 as the immunogen, with 5-fold dilution of Montanide TM GEL 01ST polymer adjuvant was mixed with equal volume, and BALB / c mice were immunized by subcutaneous multi-point injection, 3 times in total, each immunization interval was 2 weeks, and the dose was 100 μg / mice. Seven days after the third immunization, the mice were immunized with the immunogen without adjuvant. Seven days after each immunization, blood was collected to detect the antibody level by indirect ELISA. After 4 days, the mouse splenocytes with the highest titer were taken for fusion.
[0034] SP2 / 0 cells were fused with splenocytes under the action of PEG1500, and after fusion, DMEM medium (DMEM dry powder 13.5 g / L, NaHCO) was used. 3 3.7g / L, L-glutamine 0.2g / L) was added to HAT and HT for liquid excha...
Embodiment 2
[0043] Example 2 Preparation of stem cell proliferation-promoting serum-free medium
[0044] Described serum-free culture medium is made up of basal medium and the supplementary component added in basal medium, described basal medium is DMEM / F12 medium, and described supplementary component is L-glutamine, non-essential amino acid, L-glutamine. -Ascorbic acid, sodium selenite, fibronectin, ethanolamine, hydrocortisone, trypsin inhibitor, human transferrin, human insulin, bFGF, TGF-β1, PDGF-BB, coenzyme A, sodium pyruvate, and EGF and RND1-23 monoclonal antibody; wherein, the concentration of L-glutamine is 2mM, the concentration of non-essential amino acids is 2mM, the concentration of L-ascorbic acid is 50mg / L, the concentration of sodium selenite is 10μg / L, the concentration of fiber The concentration of zonulin was 20 mg / L, the concentration of ethanolamine was 2 mg / L, the concentration of hydrocortisone was 10 mg / L, the concentration of trypsin inhibitor was 1 mg / L, the co...
Embodiment 3
[0046] Example 3 Preparation of the control group of serum-free medium for promoting the proliferation of stem cells
[0047] Described serum-free culture medium is made up of basal medium and the supplementary component added in basal medium, described basal medium is DMEM / F12 medium, and described supplementary component is L-glutamine, non-essential amino acid, L-glutamine. -Ascorbic acid, sodium selenite, fibronectin, ethanolamine, hydrocortisone, trypsin inhibitor, human transferrin, human insulin, bFGF, TGF-β1, PDGF-BB, coenzyme A, sodium pyruvate, and EGF ; Among them, the concentration of L-glutamine is 2mM, the concentration of non-essential amino acids is 2mM, the concentration of L-ascorbic acid is 50mg / L, the concentration of sodium selenite is 10μg / L, the concentration of fibronectin is 20mg / L L, the concentration of ethanolamine is 2mg / L, the concentration of hydrocortisone is 10mg / L, the concentration of trypsin inhibitor is 1mg / L, the concentration of human tra...
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