69 results about "Tissue digestion" patented technology

Apparatus and methods are described for generating autologous tissue grafts, the apparatus including a point of care SVF isolation unit that includes a tissue digestion chamber in fluid communication with a lipid separating chamber, whereby SVF cells are isolated without centrifugation; and a cell seeding chamber in fluid communication with the SVF isolation unit, said cell seeding chamber adapted to support a cell scaffold. Methods and materials for cell seeding, including through the provision of micro rough scaffold surfaces, are also provided.

The invention relates to an acquisition method of adipose-derived stem cells. The method comprises the following specific steps of: digesting adipose tissues by use of mixed collagenase; culturing adipose-derived stem cells obtained through digestion in a culture solution; carrying out solution replacement and transfer of culture; and detecting to obtain the qualified adipose-derived stem cells. According to the invention, the problems that in the traditional cell separation method, cells can not be thoroughly digested by collagenase and the cell purity is low can be solved; and a separation method of adipose-derived stem cells is improved and optimized, so that tissues can be digested more thoroughly, and the purity during a culture process is higher, thus a more excellent seed cell source is ensured.

The invention belongs to the field of molecular biology and particularly relates to a kit for extracting ribonucleic acid (RNA) by virtue of a paramagnetic particle method and an extraction method. The kit contains tissue digestion fluid, lysate, proteinase K, DNase I, DNase IBuffer, nucleic acid, extraction magnetic beads, washing liquid I, washing liquid II and eluant. The invention further discloses a method for extracting the tissue RNA by virtue of the kit. According to the kit and the method, the extraction yield and purity of RNA are increased, the integrity of RNA is improved, meanwhile, the automatic extraction is realized, and the simultaneous parallel testing of multiple samples is realized, so that the labor cost and the time cost are saved.

The invention discloses placental villus plate mesenchymal stem cells. An extraction method of the placental villus plate mesenchymal stem cells includes the following steps that 1, a placental villus plate is obtained; 2, blood vessels and villus tissue are removed; 3, the villus plate is cleaned till the blood color disappears; 4, the villus plate is sheared into 0.5-5 mm<2> tissue microblocks, and the tissue microblocks are cleaned; 5, tissue digestive juice containing pancreatin, collagenase II, collagenase IV and hyaluronidase is added for digestion; 6, filtration and centrifugation are carried out; 7, culture is carried out; 8, cell digestion is carried out with 0.25% pancreatin when the cell confluence reaches 70-90%; 9, subculturing is carried out; 10, detection is carried out; 11, cryopreservation is carried out; 12, database building is carried out. According to the method, the steps are simple, and a large number of mesenchymal stem cells can be rapidly obtained from the placental tissue; the separation process does not need too much cleaning operation; the use of antibiotics is omitted, so that the purity of the placental villus plate mesenchymal stem cells obtained after separation is high.

The invention relates to a method for extracting sub-totipotent stem cells from a chorion of a fetal surface of a placenta. The method comprises the steps of removing an amnion and stagnated blood from the placenta, and then repetitively washing the surface of the placenta to perform sterilization; shearing the chorion of the fetal surface in a glass utensil, removing placenta lobule tissues remained on the surface as much as possible, and shearing the chorion into small blocks; washing small tissue blocks by using a screen and a great amount of normal saline, and removing residual blood cells; performing tissue digestion: performing oscillatory digestion in a constant temperature shaker by using mixed enzymes; adding a proper amount of FBS for termination after digestion, performing filtration through a filter screen, and adding a great amount of normal saline to wash filter residues to obtain cells as many as possible; performing centrifugation, abandoning supernatant, adding saline water for washing and performing centrifugation to obtain mononuclear cells; performing magnetic bead sorting (OCT-4 positive, Nanog positive and STRO-1 negative) to obtain target cells. The invention further relates to the sub-totipotent stem cells obtained by adopting the method and pharmaceutical use thereof. The sub-totipotent stem cells have excellent characteristics.

The invention relates to the field of organoid culture, in particular to a culture medium, a method and a kit for rapidly culturing tumor organoids. According to the culture medium for culturing the tumor organoids provided by the invention, the time for culturing the tumor organoids can be greatly shortened. The kit for rapidly culturing the tumor organoid comprises a culture medium for culturing the tumor organoid, a tissue preservation solution, a tissue digestion solution and a cell mass collection solution, and when the kit is used for culturing the tumor organoid, the success rate and survival rate of culturing the tumor organoid can be increased, an organoid model can be rapidly established within 2-5 days, which can be applied to subsequent experimental tests.

The present invention concerns a method for preparing a meat tissue digest having enhanced palatability, comprising: a) providing a meat tissue from animal; b) contacting said meat tissue with at least one alkaline endopeptidase; c) allowing said alkaline endopeptidase to hydrolyze said meat tissue; d) thermally treating the thus obtained digest product to inactivate said alkaline endopeptidase; and e) obtaining said meat tissue digest having enhanced palatability. The present invention also relates to meat tissue digests having enhanced palatability that are obtainable by such a method, and uses thereof for preparing pet food having enhanced palatability.

The invention discloses an umbilical cord tissue digestion method. According to the invention, wharton jelly is obtained from umbilical cord, twice digestion is carried out by using different tissue digestive enzyme, filtering is carried out, and the required mesenchymal stem cells can be obtained through filtering and centrifugation. The method employs twice digestion, during primary digestion, collagenase, hyaluronidase and DNA enzyme are subjected to complex formulation for usage, wharton jelly is digested, the digestive enzyme can effectively reduce the digestion consistency and has small damage on cells; pancreatin is employed during secondary digestion, and the cell separating can be more completed. The provided method can avoid the influence of long-term digestion of digestive enzyme such as collagenase and pancreatin on cell bodies, the acquisition rate of the obtained mesenchymal stem cells is high, and the obtained mesenchymal stem cells have high activity and differentiation potential.

The invention discloses a separation method of human placenta mesenchymal stem cells. The separation method comprises the following steps that placenta specimen treatment is carried out, treated placenta tissue is obtained, tissue digestion is carried out on the placenta tissue, and cell suspension is obtained; preliminary subculture of the placenta mesenchymal stem cells is carried out; further separation and culture of the placenta mesenchymal stem cells are carried out. HyQTase and DNAse I are adopted for digestion together, the placenta mesenchymal stem cells are obtained through separation, after preliminary culture, microgravity treatment and electromagnetic field and sound wave treatment are carried out, BMP4 is used for treatment many times, upper layers are removed through a differential attachment method, a serum-free medium and antioxidant are replenished in a culture bottle for culture, and the final purity is high. The stem cell characteristics of the cells are verified through detection on hereditary stability, cell surface molecule expression conditions and cellular morphology in the continuous passage process, and materials are provided for seed cells with the placenta mesenchymal stem cells as clinical application.

The invention discloses a human adipose sub-totipotent stem cell isolated-culture method and a stem cell bank establishing method. The human adipose sub-totipotent stem cell isolated-culture method includes the following steps of human adipose tissue collection, sub-totipotent stem cell separation and obtaining, sub-totipotent stem cell culture and proliferation, sub-totipotent stem cell cryopreservation, product quality control, stem cell bank establishing and stem cell revivifying and annual inspection. Mixed collagenase used by the method can effectively separate sub-totipotent stem cells from adipose tissues, is homogenous in tissue digestion, no obvious tissue residual is produced, the cell activity is good, wall adherence is quick, and the homogeneity is good. Cryopreservation liquid adopted by the method can effectively maintain the cell activity, a gradient cooling box is adopted to cryopreserve cells, the cell activity is not affected, and the human adipose sub-totipotent stem cell isolated-culture method is simple in operation, convenient and high in efficiency.

The invention provides a separation and enrichment method for quickly extracting tissue extracellular vesicles. The separation and enrichment method comprises the following steps of: A: shredding target tissues by a mechanical method, adding tissue digestive enzymes to carry out tissue dissociation, and filtering obtained tissue dissociation liquid to obtain a tissue cell suspension; and B: carrying out differential centrifugation, ultracentrifugation, SEC exclusion and ultrafiltration on the tissue cell suspension in sequence, and carrying out enrichment and purification of the tissue extracellular vesicles. The tissue extracellular vesicle separation and enrichment method provided by the invention greatly simplifies a step of separating the tissue extracellular vesicles, enrichment timeis saved, and a whole process only needs 4-5h. The enriched extracellular vesicle has high purity, small soluble impure protein pollution and wide practicability. The extracellular vesicle enriched bya tiny quantity of tissue samples can meet subsequent analysis, including NTA, WB, electron microscopes, transcriptomes and the like.

The present invention provides a method for promoting directional differentiation of umbilical cord mesenchymal stem cells into chondrogenesis. The method comprises the following steps: taking out umbilical cord tissues and conducting shredding and washing, adding tissue digestion fluid for digestion, and conducting filtering to remove supernatant after the digestion to obtain umbilical cord mesenchymal stem cells; culturing the umbilical cord mesenchymal stem cells with an umbilical cord mesenchymal stem cell selective culture medium; when the cells reach 80-90% confluence, digesting the cells with low-toxic digestion fluid, then using an umbilical cord mesenchymal stem cell cryopreservation solution for cryopreservation and placing the cells in liquid nitrogen; removing the cells from the liquid nitrogen, conducting quick thawing in a 37-DEG C water bath kettle, conducting centrifugation to remove supernatant, and adding a cell recovery solution to recover the cells; removing the recovery solution and replacing an exciting solution before induction; and when the cells grow to 80-90% confluence, digestign the cells with the low-toxic digestion solution; inoculating the cells, conducting culture overnight, removing the supernatant, adding a chondrogenesis induction culture medium, and conducting culture in a hypoxic incubator at 2% oxygen concentration. Compared with the priorart, the method can stably and effectively promote the directional differentiation of the umbilical cord mesenchymal stem cells into the chondrogenesis with shorter induction time.

The invention discloses a preparation method of a fish tissue single cell suspension. The preparation method comprises the following steps of: 1) preparing a reagent and a culture medium; 2) pretreating tissues; 3) performing tissue digestion and dissociation; 4) terminating digestion; 5) filtering by using a micron cell filter; 6) obtaining a target tissue single cell suspension; and 7) measuring the yield of living cells. The preparation method of the fish tissue single cell suspension can be used for quickly and efficiently obtaining the fish tissue single cell suspension at low cost, ensures that dissociated cells keep good survival rate, can be applied to dissociation, separation and purification of large-scale fish cells and establishment of cell lines, and provides important reference data for research of fish single cells, gene functions and genetic breeding.

The invention relates to a method for separating scylla paramamosain tissue exosome. The method mainly comprises the following steps: S1) performing tissue extraction; S2) performing tissue digestion;S3) performing low-speed centrifugation; S4) performing low-speed centrifugation again; S5) performing high-speed centrifugation; S6) performing high-speed centrifugation again; S7) resuspending andprecipitating; S8) performing sucrose density gradient centrifugation; S9) carrying out ultracentrifugation; S10) carrying out ultracentrifugation again; S11) dialyzing in a dialysis bag; S12) filtering with a filter membrane; S13) carrying out ultracentrifugation; and S14) resuspending and precipitating to obtain the extracted and separated exosome product. According to the method for extractingthe scylla paramamosain tissue exosome, the differential centrifugation method, the sucrose density gradient centrifugation method and the dialysis filtration method are combined, the method is stable, effective, high in purity and easy to operate, no special equipment is needed, and the extracted exosome is high in content and purity.

The invention discloses a preparation method of a mouse meningeal single-cell suspension. The preparation method comprises the following steps of: 1) preparing a reagent and a culture medium; 2) carrying out tissue pretreatment; 3) carrying out tissue digestion and dissociation; 4) terminating digestion; 5) filtering by a micron cell filter; 6) obtaining a target tissue single-cell suspension; and 7) measuring the yield of living cells. The preparation method of the mouse meningeal single-cell suspension can be used for quickly and efficiently obtaining the mouse meningeal single-cell suspension at low cost and ensuring that dissociated cells keep good activity, can be applied to dissociation, separation and purification of mouse meningeal cells in a large range and establishment of a cell line, and provides important reference data for research of mouse nerve tissue single cells, gene functions and genetic breeding.

The invention relates to a mesenchymal stem cell separated from the placenta blood vessel with a digestive enzyme composition, in particular to the digestive enzyme composite used for a method for separating the placenta mesenchymal stem cell from the placenta blood vessel. A buffering solution containing tissue digestive enzymes is provided with added digestive enzymes which include pancreatic enzymes, deoxyribonuclease I, collagenase II, collagenase IV and hyaluronidase. In addition, the invention further relates to the method for separating the mesenchymal stem cell from the placenta bloodvessel, and the method includes the steps that the placenta is sterilized; the placenta vessel is separated from the placenta; cutting, cleaning and bloodiness filtering removal are conducted, so thatplacenta blood vessel tissue is obtained; mixed enzyme liquor is added for digestion; digestion is stopped, and interstitial fluid is filtered and collected; cell sediments obtained through centrifuging are original placenta mesenchymal stem cells, re-suspending is conducted, sampling is conducted, and the number nucleated cells and the survival rate of the nucleated cells are calculated; the obtained cells are subjected to refrigeration preservation or continuous passage and / or identified, detected and subjected to refrigeration preservation and database creation. Through the method, the efficiency of separating the mesenchymal stem cell from the placenta blood vessel can be improved effectively.

The invention relates to a method for measuring the digestive enzyme activity of tissues of a seahorse baby with enzyme linked immunosorbent assay. The method comprises the following steps of: (1) putting a frozen seahorse baby into a container, adding normal saline of NAC1 with the concentration of 0.7 percent or a homogenizing medium in an ice bath, homogenizing by using a multifunctional sample homogenizer, centrifuging with a centrifuge and taking supernatant fluid for later use; (2) measuring the total protein concentration of the tissues with a part of the supernatant fluid; (3) adding a substrate starch buffer solution into a container A and a container B respectively, adding the supernatant fluid obtained in the step (1) into the container B and adding iodine reaction liquid and distilled water into the containers A and B respectively; (4) sucking the solutions in the containers A and B, dripping the solutions into two holes of an enzyme label plate, putting the enzyme label plate into an enzyme label reader and detecting the absorbency of the solutions in the two holes simultaneously; and (5) calculating the amylase activity of a sample. The method disclosed by the invention is easy to operate, has low cost and high efficiency and is environment-friendly.

The invention discloses a preparation method and a resuscitation method of decidua parietalis mesenchymal stem cells, and the preparation method comprises the following steps that tissue blocks are digested by using a high-glucose DMEM culture medium containing 40-60% by volume of Tryple-EDTA enzyme and 8-12 mg / ml of type II collagenase as a tissue digestion solution, so that the decidua parietalis mesenchymal stem cells can climb out of tissues for adherent growth; a DMEM (dulbecco's modified eagle medium) serum-free culture medium containing a serum substitute with the volume concentration of 8-12%, L-glutamine with the volume concentration of 0.5-1 mol / ml, a basic fibroblast growth factor with the volume concentration of 18-25 ng / ml, an epidermal growth factor with the volume concentration of 16-22 ng / ml and a stem cell growth factor with the volume concentration of 6-12 ng / ml is adopted as a selective culture medium to terminate digestion, and the decidua parietalis mesenchymal stem cells are resuspended, so that the purity of the decidua parietalis mesenchymal stem cells is improved, the growth of the decidua parietalis mesenchymal stem cells is accelerated, and the in-vitro rapid amplification of the decidua parietalis mesenchymal stem cells is realized.

A tissue digester system includes a container for housing a digestion chamber having an exterior vessel for holding digestor fluid and an interior vessel, the container extending from a first end to a second end, the interior vessel having perforations and having baffles extending from an interior surface of the interior vessel; a lid secured to the exterior vessel and to provide access to the digestion chamber; one or more heating elements positioned to apply heat to the digestion chamber; a motor engaged with the interior vessel and to create rotational movement of the interior vessel; a control system, having a temperature controller; and a movement controller; the control system is to rotate the interior vessel and heat the digestion chamber based on user commands; and the digestion chamber is to break down remains through application of the digestor fluid to the tissue remains.

To enable to yield a group of fat-derived stem cells of constant quality regardless of the individual difference in fat tissue. It is intended to provide an apparatus for determining the termination of fat digestion (1) for use in a fat tissue digestion apparatus which digests a fat tissue by placing and stirring the fat tissue with an enzyme-containing physiological saline, lactate Ringer's solution, or buffer solution in a container, wherein the apparatus for determining the termination of fat digestion comprises: a boundary-detecting unit (8) which detects the boundary between a fat tissue layer formed in the container by being left still after stirring and a cell suspension layer located below the fat tissue layer, from outside the container; a layer thickness-measuring unit (10) which measures the thickness of the fat tissue layer based on the position of the boundary detected by the boundary-detecting unit (8); a fat volume-calculating unit (12) which calculates the volume of the fat tissue layer by multiplying the thus measured thickness of the fat tissue layer by the cross-sectional area of the container; and a termination-determining unit (13) which determines the termination of digestion based on the thus calculated volume of the fat tissue layer 120

The invention relates to an in vitro culture method of limbal stem cell stability. The method is characterized in that after treated by a gentamicin solution, the limbal tissue is cut into small blocks, and then placed in a tissue digestion solution for digestion and filtration, and tissue cell precipitate is collected, and then the precipitate is resuspended by a primary culture solution of limbal stem cells and is inoculated to a cell culture plate pretreated with a limbal stem cell coating solution for primary culture, after a cell cloned sphere appears, the cell cloned sphere is digested with the tissue digestion solution, then centrifugation is performed, the cell precipitate is collected, and the cells are resuspended by a subculture solution of the limbal stem cells, then are inoculated to the pretreated cell culture plate for subculture to obtain stable limbal stem cells. In order to obtain more stable limbal stem cells, multi-generation expansion culture can be carried out after the subculture, and a large number of stem cells with good stability and high safety can be obtained so as to meet the requirement of clinical application.

The invention relates to a separation and purification method of umbilical cord mesenchymal stem cells and belongs to the technical field of stem cells and regenerative medicine. The method comprisesthe steps as follows: separation of umbilical cord tissue: taking the umbilical cord tissue, cleaning the umbilical cord tissue and removing blood, and shearing the tissue into tissue fragments for later use; tissue digestion: taking the tissue fragments, and adding tissue digestive juice for oscillation digestion, wherein the tissue digestive juice contains neutral protease, hyaluronidase, collagenase II and DNA enzyme; after digestion, adding a stop buffer for stop, performing filtration to obtain undigested tissue blocks and filtrate, and centrifuging the filtrate to remove supernatant to obtain cells; and cell culture: inoculating a culture flask with the tissue blocks and cells, using serum-free culture medium for culture to obtain the umbilical cord mesenchymal stem cells. The separation and purification method of umbilical cord mesenchymal stem cells overcomes the defects of an enzyme digestion method and a tissue block culture method in the prior art and can obtain enough primary cells within 3-5 days.

The invention provides a separation method of human amniotic mesenchymal stem cells. The separation method comprises the following steps: (1) collecting human placental tissues and separating amniotictissues; washing blood stains; (2) chopping the amniotic tissues and adding a first tissue digestion solution and carrying out digestion; (3) after finishing the digestion, removing the first tissuedigestion solution; washing the amniotic tissues; adding a second tissue digestion solution and carrying out digestion; (4) after finishing the digestion, adding a trypsin solution and carrying out digestion; (5) adding a stop solution and stopping the digestion; filtering to remove tissue blocks which are completely digested, collecting a cell suspension solution and centrifuging; (6) removing supernatant, adding a DMEM (Dulbecco's Modified Eagle Medium) and washing sediment; centrifuging again to obtain the separated human amniotic mesenchymal stem cells. The method provided by the inventionis simple and effective, can obtain high-purity human amniotic mesenchymal stem cells and has no influences on cell activity.

This invention discloses a digest and separator for biology apparatus organization, which sets a temperature detector on a drum connected with a cover by screws, a mesh and a sealing loop are set between the drum and the cover for filtering, a filter hole is set at the bottom of the frame of the mesh and a clamping ring clamps the mesh on the frame.

Tissue digestion methods and apparatuses that provide easy, safe and inexpensive disposal of biological tissue, for example animal carcasses and human cadavers, are disclosed. Embodiments include tissue digesters with elongated cylindrical vessels for holding digestive fluid and baskets for holding tissue within said vessel. Embodiments include baskets with perforations to allow circulation of digestive fluid around and about the tissue. In one form the basket holding the tissue is horizontally loaded into a horizontally disposed vessel. The vessel is then tilted to a more vertical orientation. Gravity helps to collect the tissue fragments in a sloped collection region of the basket, which is located near a mixer to allow continual agitation of the tissue fragments as they are digested. The tissue digester can operate efficiently at lower temperature and pressures, is mechanically less complicated, consumes less power and is less expensive to manufacture than conventional tissue digesting systems. At higher temperatures employed in a pressure vessel provided by this invention, the tilting mechanism also dramatically improves the efficiency of the process and the completeness of the tissue digestion.

The invention relates to the technical field of cell separation, in particular to adipose tissue digestive juice and a method for rapidly obtaining an SVF (stromal vascular fraction) cell. The adiposetissue digestive juice comprises the following ingredients: I-type collagenase, HEPES buffer solution and a DEMEM / F12 culture medium. According to the adipose tissue digestive juice provided by the invention, the pH value of a digestive system can be stably maintained between 6.8-8.2, so that the activity of the I-type collagenase is optimal. Therefore, complete digestion can be realized in a short time. The time for obtaining the SVF according to the traditional digestive scheme is 40 min, while the SVF with high quality and high quantity can be obtained within 15-20 min in such a way that the digestive juice is adopted for digesting adipose tissues.

The invention provides a method for preparing dental pulp mesenchymal stem cells. The method comprises the following steps: adding tissue digestion liquid into dental pulp tissues for digestion; afterthe digestion is finished, adding H-DMEM culture liquid to dilute digestion liquid, filtering tooth fragments, and centrifuging the filtrate to remove supernate, so as to obtain precipitates; cleaning the precipitates with the H-DMEM culture liquid, and carrying out centrifugation again to remove the supernate, so as to obtain the separated dental pulp mesenchymal stem cells. According to the method, the dental pulp mesenchymal stem cells can be effectively prepared, separated and purified.

The invention relates to a method for preparing amniotic mesenchymal stem cells from a human placenta amnion and application of the amniotic mesenchymal stem cells. The method comprises the following steps: washing and disinfecting a placenta; washing and disinfecting an amnion and crushing the amnion; carrying out tissue digestion; carrying out cell culture on a filtrate part and a tissue block part; harvesting the cells; counting and determining the survival rate of the cells, and cryopreserving. The invention further relates to the amniotic mesenchymal stem cells prepared by the method and therapeutic application of the amniotic mesenchymal stem cells. The method provided by the invention has the advantages showed in the description.