7 results about "Heparinase" patented technology

The invention belongs to the technical field of bioengineering, and relates to heparan sulfate defect type matrigel as well as a preparation method and application thereof. The preparation method comprises the following steps: providing natural matrigel; providing immobilized heparinase, wherein the immobilized heparinase can specifically catalyze and degrade heparan sulfate in the natural matrigel; carrying out contact reaction on the natural matrigel and the immobilized heparinase; and after the reaction is finished, separating the immobilized heparinase. According to the preparation method, the characteristics that the immobilized enzyme is convenient to separate and recycle are utilized, the problems that in a traditional solution enzyme method, enzyme residues exist, the reaction is difficult to accurately control, and subsequent purification steps are complex are solved, and a standardized heparan sulfate defective matrigel product which is controllable in component, free of residual enzyme and high in stability is provided. The method has wide application value in the fields of tumor mechanism research, anti-tumor drug screening, tissue engineering and the like.

The application belongs to the technical field of heparin extraction, and particularly relates to a method for preparing low-molecular heparan sulfate by using an enzyme method. The method comprises the following steps: (1) dissolving heparan sulfate in deionized water, adjusting the pH to 8-8.5 by using a NaOH solution at 20-25 DEG C, adding modified carbon nanotubes loaded with heparinase, then performing enzymolysis for 3.0-3.5 h under stirring at 35-45 DEG C, and obtaining an enzymolysis solution; (2) when the specific absorbance of the enzymolysis solution in step (1) is 1.3-1.5 at a wavelength of 230 nm, rapidly heating the enzymolysis solution to 95-98 DEG C, and terminating the reaction after 0.5-1.0 h; and (3) cooling the enzymolysis solution after the reaction to 20-30 DEG C, adding anhydrous ethanol, stirring for 1-2 h, centrifuging to obtain a precipitate, adding deionized water, filtering by using a 0.1-0.22 mu m filter membrane, and vacuum freeze-drying to obtain low-molecular heparan sulfate. The application has a mild reaction and a high yield.

The invention provides a construction method of a heparinase fingerprint spectrum, which comprises the following steps: a) taking a heparinase sample to be detected and diluting with a diluent to prepare a heparinase test solution with a certain concentration; and b) under normal pressure, determining the retention time and peak area A enzyme of each enzyme in the heparinase test sample solution by adopting a high performance liquid chromatography analysis method, and recording a spectrogram. The fingerprint spectrum of the heparinase and the construction method of the fingerprint spectrum can be widely applied to quality detection and control of a crude heparinase product and a finished heparinase product in the preparation process of the heparinase, and the method for detecting the quality of the heparinase by utilizing the fingerprint spectrum of the heparinase is simple, convenient, rapid and accurate to operate.

The invention belongs to the technical field of immune cell therapy, and particularly relates to a heparanase-loaded CAR-T exosome and application thereof. According to the invention, the HPSE-loaded exosome is secreted by the engineered CAR-T cell, and the activity of the HPSE is limited in a tumor microenvironment (TME) by utilizing an anchoring mechanism, so that systematic off-target toxicity is avoided. The result of the embodiment shows that the exosome shows remarkable ECM degradation capacity, tumor killing efficiency and immune regulation function in in-vitro and in-vivo models, has the characteristics of simplicity in preparation, high safety and remarkable curative effect, and provides a new solution for solid tumor immunotherapy.

The invention provides a large-scale multiplication culture method of CAR-T cells, and belongs to the technical field of CAR-T cells. The preparation method comprises the following steps: coupling bevacizumab and heparinase, immobilizing the coupled bevacizumab and heparinase on anti-CD3 / CD28 magnetic beads to prepare modified magnetic beads, resuspending CAR-T cells by using an MACS T cell culture medium containing potassium chloride, decitabine liposome and 2-deoxy-D-glucose, adding the modified magnetic beads, uniformly mixing, and performing magnetic field stimulation culture to obtain the high-activity CAR-T cells. The high-activity CAR-T cells prepared by adopting the large-scale amplification culture method of the CAR-T cells not only have better wettability, can enter a tumor microenvironment, but also can improve recruitment of the CAR-T cells in the tumor microenvironment, and have better anti-tumor activity, meanwhile, the amplification rate is obviously improved by adopting the method, and the method is suitable for large-scale in-vitro amplification culture.

The present invention is directed to synthetic hexasaccharide mimics of heparin having formula (I) wherein: R1 is selected from the group consisting of NH2, NHSO3 -, NHAc; R2 is selected from the group consisting of H, SO3 -, Bn, 2-Naphthylmethyl, preferably H; R3 is selected from the group consisting of H, Ac, SO3 -; R4 is CO2 -; R5 is Me, Et, C3-C30 linear or branched alkyl, optionally comprising one or more isolated or fused rings, PEG-OMe; R6 is selected from the group consisting of H, Ac, SO3 -; R is a C10-C30 linear or branched alkyl optionally comprising one or more isolated or fused rings. In another embodiment, the invention is directed to the use as a drug of hexasaccharides having the above formula, wherein R, R1, R2, R3, R4 and R6 have the above defined meaning, while R5 is selected from the group consisting of Me, Et, C3-C30 linear or branched alkyl, optionally comprising isolated or fused rings, PEG-OMe.