CN110368526A - A kind of medical acellular organism material and preparation method thereof - Google Patents

Abstract

The invention belongs to medical repair materials fields, are related to one kind for tissue repair and remove cell material and preparation method thereof with medical.Of the invention goes cell material and preparation method, both without using any chemical cross-linking agent or optical means, also without making up the problem that current material stability is poor, degradable, mechanical property is insufficient using physics class method, chemical agent residue and toxicity are avoided.The present invention picks the connective tissue of breeding stock as raw material, does not select commodity livestock raised for meat as the source of raw material, and the connective tissue of breeding stock has stronger toughness, better biomechanical property than commodity livestock raised for meat.Of the invention removes cell medical material, existing the advantages of removing cell material can not only be retained, also there is better stability and mechanical property, it is not easy to break, it is easy to suture and good leak tightness, is particularly suitable for the tissue repair of the tissue site high to stability and mechanical property requirements.

Description

A kind of medical acellular organism material and preparation method thereof

Technical field

The invention belongs to biomaterial for medical purpose field, it is related to a kind of medical acellular organism material and preparation method thereof.

Background technique

The biomaterial used on clinical medicine at present is divided into synthesis class material and natural biologic material two major classes；Synthesis Class material, mechanical strength is preferable, and stability is good, not degradable, but is unable to inducing tissue regeneration and healing.And natural biological class Material；Mostly remove cell material, be mainly derived from mammal corium, small intestine, pericardium etc. tissue, by degreasing, go cell, The process such as DNA are gone, to obtain the three-dimensional structure for remaining with extracellular matrix and some active ingredients, including collagen The ingredients such as albumen, non-collagen and growth factor provide the environment of adaptation for host cell attachment, proliferation, differentiation；Facilitate The structural remodeling of damaged tissues, and then the reparation and regeneration and functionally rehabilitation of promotion tissue.

The main problem currently encountered is, this kind of to remove cell medical material, in order to ensure highly safe, it is ensured that do not have Immunological rejection, the danger not being infected；In practical application production, in this kind of production technology for removing cell material, Cell link is especially gone, it is all very harsh, and it is all longer to handle the time, sometimes up to 10--20 more a hours, this place The reason of reason has different degrees of influence and destruction to tissue ECM, organizes raw material itself in addition, causes existing to remove the medical material of cell Material, stability is poor, degrades fast, and sheet finished product (referred to as biological sticking patch) mechanical strength is insufficient；In order to make up this respect not Foot, overcomes defect existing for finished product；Sometimes in the process of this kind of medical material, epoxides or penta are additionally used The chemical cross-linking agents such as dialdehyde, or shared with synthetic material, it is just able to satisfy requirement of the finished product in mechanical property and prevents from degrading Fastly.But after chemically crosslinking agent, it is understood that there may be following disadvantage: having potential cytotoxicity, and the degree of cross linking is difficult to accurate control； If excessive crosslinking can degradation rate it is slower, mismatched with regeneration, can lead to the reaction such as fibrosis, chronic inflammation.

Application No. is the Chinese patent of 201710862130.X, right-angled intersection ligament reproducibility implantation materials and preparation method thereof With application, although reaching the mechanical requirements of tissue repair operation, but by degradable high polymer and fibrinogen After being blended Deng the biomaterial with guide tissue regeneration, it is prepared using electrospinning technology.

Application No. is the Chinese patent of 201310203598.X, a kind of bioartificial tissue and preparation method thereof is only simple Single curling or superposition, the active force very little between each layer, it is likely that lead to the inhomogenous and each layer of stress layer because of layering The case where rupturing respectively.

Ideal goes cell medical material that should have following condition: 1. having safety, non-toxic, 2. no infectivity is organized Compatibility is good, no immunological rejection.3. compactness is good, no permeability.4. if then there is toughness for suturing, it is easily firm Gu suture, not easy to break.5. regeneration can be promoted, do not stick together.6. it is easy to use, simply, it is easy to sterilization.7. taking Material is extensive, cheap.

About the mechanical property for removing cell medical material, clinically mostly using corium, pericardium, the abdomen for being originated from commodity livestock raised for meat Film, tissue, small intestine etc. are as the initial raw material for removing cell medical material；In practical application, this kind of sticking patch is normal in mechanical property The mechanical requirements of cell medical material are not achieved；Therefore during cell medical material is removed in preparation, frequently with following two kinds Method reaches the high requirement of good toughness, tensile strength come the stability for enhancing sticking patch mechanical property with enhancing product.

First is that: chemical (biochemistry) method uses epoxides, glutaraldehyde, Geniposide, paddy ammonia in sticking patch preparation process All kinds of crosslinking agents such as amide transaminase (TGase), to improve the biomechanics characteristic of sticking patch.The disadvantage is that crosslinking degree is difficult to control System, may there is chemically residual crosslinker sometimes, these crosslinking agents have potential cytotoxicity, and this kind of cross-linking type sticking patch Usual mechanical property is good, and resistance is strong, and slowly, degradation is unable to simultaneously match with regeneration for not degradable or degradation, easily leads to The adverse reactions such as fibrosis, chronic inflammation；This kind of sticking patch mechanical strength is enough simultaneously, but it is slower to degrade, and can lead to fibrosis, chronic Inflammatory reaction；And because its is harder, can not be bonded with tissue surface perfection.

Second is that: physical method, such as laterally or longitudinally by single-layer sheet, the overlapping of multilayer cross section are placed, although adopting on the whole The sticking patch being prepared into the method, tensile strength increase, but more using sticking patch raw material, are on the one hand that raw material is unrestrained Take, be on the other hand that sticking patch is blocked up, the time that entire sticking patch can be made degradable is dragged too long, and then influences repairing for target tissue Multiple and function rehabilitation.In addition sticking patch raw material is made after fine strip shape and is twisted line, then sticking patch is made by way of braiding；By It weaving and manufactured sticking patch, drawing stretches intensity and is improved significantly, but first slice is made in sticking patch raw material by this step needs, then It is prepared by weaving manner, technically there is a difficulty, it is a little complicated in step.

Summary of the invention

One of it is an object of the invention to, it is that solving part removes cell medical material, in actual clinical in use, stablizing Property it is poor, be easy degradation, or degradation is very fast, cannot effectively meet and be actually needed on clinical medicine, such as product degradation is too fast, it is unfavorable In assisting the generation of damage location cambium and the rehabilitation of function as reliable and stable cytoskeleton.

It is an object of the invention to it two, be solve part remove cell medical material, in actual clinical in use, mechanics Performance is poor, weaker, and rupture, tearing or other accidents easily occurs in when actual clinical use, cannot effectively meet clinical medicine reality Border needs, such as hernia sticking patch, tendon sticking patch, rotator cuff sticking patch and dural patch of poor mechanical property etc. will appear spalling, drawing crack Or crack.

It is an object of the invention to it three, be to avoid crosslinking agent that may be present in existing medical material, weaving method Method and product in cell material stability and mechanical characteristics, existing aforementioned various insufficient and defects are gone in equal enhancings.

Inventor is read around the stability and biomethanics these specified features of material by lot of documents, is tied Itself is closed to the further investigation of academic theory and is carefully analyzed, in addition working experience abundant for many years, close by theory and practice In conjunction with ingenious and insufficient, the weak technical problem of mechanical property that easily solves cell medical material stability.

Main innovation point of the present invention is, the tissue or organ for selecting breeding stock are as the raw material for removing cell, rather than commodity Livestock raised for meat.

Because existing animal sources go cell materials to be taken from fresh slaughtered animals, gone cell technique and etc. processing and At, used fresh slaughtered animals tissue, and be all the pork pig, beef cattle, mutton sheep for the commercialization just delivered for sale；Such as slaughter in pig farm to It kills, the overwhelming majority is the commercialization pork pig just delivered for sale, and three way cross in addition, such as based on Ternary Pig kind, also there is one Ternary pork pig kind in a little；The weight of pork pig is delivered for sale in 90-120 kgs；Raising age in days is 5-6 months, by season and pig price Market, pork pig delivers weight for sale and number of days is changed.

Inventor solves to go poor, degradable, product the mechanical property of stability present in cell medical material weak etc. to ask Which kind of mode weight is the technical thought of topic such as use neither resting on the subtle improvement for processing physical method when removing cell material Folded (vertical/horizontal/tiltedly), overlap ratio be how many, it is several layers of to need to be overlapped, is used when weaving raw material line thickness, be plain weave knit, twill weave Or the modes such as knit are twisted, nor resting on the variety selection of crosslinking agent, proportion, concentration are improved, the technologies such as crosslinking time length The optimization of parameter；But picked up from the source for preparing the i.e. raw material of the acellular organism material first step, from initial animal sources, It is furtherd investigate as technological break-through point or impetus, the main innovation point of product of the present invention, in that remove cell material The selection of animal tissue's raw material of material, the not now commerical grade livestock raised for meat that everybody uses, but select the prime of life breeding stock, preferably in Prime of life Suprapubic arch sling and kind cow, preferably healthy eliminates dam because of reproductive performance decline.

Inventor is in deep pig raising First Line, after sufficiently linking up with large-scale pig farm responsible person, is supported according to what it was told about It grows mode (being usually all home-bred and autophytic) and the culture-cycle (delivers for sale, about 90-120 kg of weight for pork pig 5--6 months；And sow It after eight to ten tires, is eliminated gradually because of reproductive performance decline, raises existing 36 wheat harvesting periods at this time, weight is up to 180 kilograms) Actual conditions, inventor by taking respectively, and the tissue of the different parts of sow and Market pigs is (under corium, mucous membrane of small intestine Layer), through substantially similar material preparation method (including going cell technique etc. using identical) preparation finished-product material；Then distinguish The mechanical property of these finished-product materials is detected, discovery uses the various types of materials in sow source, and tensile strength (i.e. toughness) will compare Market pigs source material with position is significantly much higher；In addition select the organ or tissue from superseded sow as preparation Medical acellular organism material, also has obviously price advantage, and the price of identical tissue site eliminates sow, all It is lower than Market pigs, such as same secondary small intestine, the price for eliminating sow want low 30-50%；Selection, which is originated from, eliminates sow Respective organization position such as small intestine, corium etc. is really super quality and competitive price as the raw material for removing cell；It kills two birds with one stone.

The secondary innovative point of product of the present invention is that preferably tissue site is submucous layer of small intestine (SIS), also can choose Corium；Because this class loading removes cell and manufactured material, all containing natural three-dimensional three-dimensional structure, but with other tissue sites Comparatively, the composition of submucous layer of small intestine (SIS) is constituted based on I fiber type collagen, while also containing III, IV, VI Collagen type especially wherein contains important IV collagen type, has apparent promote to the formation of new blood vessel and basement membrane Into effect；In addition the plurality of articles delivered according to U.S. STEPHEN F. doctor BADYLAK show the catabolite of SIS sticking patch With stronger antibacterial activity, there is the function similar to porcine defensin (porcine defensin, pBD-1)；And George S.Hussey article in 2018 " Extracellular Matrix Bioscaffolds for Building Gastrointestinal Tissue " report that SIS sticking patch catabolite also has induced cellular chemotaxis and mitotic work With.

Inventive principle:

Of the invention removes cell medical material, and de-cellular system raw material sources are breeding stocks, such as have produced the sow of several tires, body Weight usually significantly weighs than Market pigs, and volume is also obvious big；Some adult Suprapubic arch slings are up to 200 kilograms, raising Duration is usually more than 24 months, its raising age in days of superseded sow is likely to be breached 40 months or more long, weight is even more to be more than It is 200 kilograms, a other to reach 220-250 kilograms.Relative to Market pigs, the tissue and organ of adult sow various aspects are sent out It educates all very completely, realizes real meaning in sub- organ level and structure, the indexs such as sight and micro-property, histology intensity that are situated between On, it is sufficiently and comprehensively mature；Further from molecular level and molecular structure, possible collagen degree of hydroxylation Height, triple-helix structure stability is more preferable, and the natural degree of cross linking is high between collagen, and stability is preferable, not degradable；Usual collagen Cross-bond is amino and the aldehyde radical by lysine or hydroxylysine, and under the action of lysyloxidase, it is anti-to carry out oxidative deamination It should be formed, the covalent bond of gained lysine formula is highly stable；It is preferable in the mechanical property for macroscopically showing as connective tissue, system It is standby at material, the tensile strength that detected is high.

And it is present remove cell material, raw material sources are all commodity livestock raised for meats, such as pork pig or beef cattle, usually raising week Phase is short；Such as pork pig breeding cycle only has 5-6 months, weight when delivering sale for sale is about 90-120 kgs, relative to Weight 180--200 of the numerous sow of energy or so or heavier (presses kind and parity, the weight of sow is different), is obvious partially light , the only half or so of its weight；Each histoorgan of commodity livestock raised for meat simultaneously, relatively for the numerous sow of energy, only at the beginning of surface Tool scale, frame structure forming, but actually there is no really thorough for the indexs such as its internal structure, micro-property, histology intensity Bottom it is sufficiently and comprehensively mature, further from molecular level and molecular structure, possible collagen degree of hydroxylation Low, triple-helix structure stability is poor, and the natural degree of cross linking is low between collagen, and stability is poor, is relatively easily degraded；In macroscopically table Now poor for the mechanical property of connective tissue, the material being prepared into, the tensile strength that detected is low.

Compared with prior art, the present invention have following remarkable advantage and the utility model has the advantages that

1. cell medical material product is removed in invention, using breeding stock tissue as raw material, there is the good natural degree of cross linking, stability is good, Not degradable, manufactured material has good toughness, tensile strength height, is not easily broken, and wear-resisting property is good；

2. invention goes cell medical material product to remain in extracellular matrix with submucous layer of small intestine, corium etc. for raw material The ingredients such as stereoscopic three-dimensional structure, types of functionality albumen, growth factor and glycosaminoglycan have good inducing tissue regeneration function Can, the growth of postoperative tissue tissue and the reconstruction of function can be accelerated；

3. cell medical material product is removed in invention, there is no crosslinking agent in production, will not crosslinking dose residual, without potential cell toxicant Property, it not will lead to fibrosis, chronic inflammation；

4. cell medical material product is removed in invention, degradation speed is suitable, too fast will not degrade, degradation speed can be more slowly；Simultaneously Its ECM three-dimensional structure tool inducing cell and blood vessel grow into function, itself can gradually degrade while new tissue is grown into.

In addition from economics, raw material can be the Suprapubic arch sling that is eliminated due to reproductive performance decline, usually this Class cost of material will be significantly lower than Market pigs；For in high volume to purchase for raw material in industrialized production from now on, use Sow is eliminated as the raw material for removing cell, the spending of writing can be saved for enterprise, this is also one of innovative point.

The object of the present invention is to provide one kind to remove cell medical material and preparation method thereof, to realize mesh of the invention , it is realized particular by following technical scheme.

Described to remove cell medical material, tissue raw material is derived from adult breeding stock, rather than is originated from commodity livestock raised for meat.

Further, it is described adult breeding stock refer to adult and raising purpose be mainly used for the kind used that raises up seed Pig, kind ox, the medium-and-large-sized mammal for planting sheep, horse, camel, donkey, mule and other artificial feedings through producing.

Further, the adult breeding stock refer to sow through producing, cow, ewe, mare, female camel, jenny ass, female mule and The medium-and-large-sized mammal of other artificial feedings through producing.

Further, preferably adult breeding stock is sow, cow, ewe, the mare, mother being eliminated by reproductive performance decline Camel, jenny ass, female mule etc..

Further, the raw material for removing cell medical material include the submucous layer of small intestine of breeding stock, corium, internal organs film, Peritonaeum, pericardium one or more raw materials combination.

Further, the raw material for removing cell medical material is the submucous layer of small intestine and corium for eliminating sow, cow.

Further, it is described go cell medical material be it is preprocessed, disinfection, degreasing, remove cell, remove DNA and remove α-Gal The techniques such as antigen and dry sterilization are process；Also may be selected by finished product target physical state, using physics, chemistry or The method of biological (such as enzyme) is processed into corresponding state, such as solid sheet, graininess, gel, then carries out sterilization treatment.

The present invention also provides a kind of preparation method for removing cell medical material, main includes the selection of first step raw material, Select the respective organization position of breeding stock as the raw material for removing cell, the preparation method of specific product includes:

1) it draws materials and cleans: taking the connective tissue of breeding stock, sufficiently clean；

2) it pre-processes, three pre-sterilization, degreasing links are handled；

3) cell is removed: using surfactant, protease, EDTA or other solution or mixed solution for going cell reagent to be made into, Under ultrasound condition, cell materials are removed in immersion, are rinsed later with PBS；

4) it removes DNA and removes α-Gal antigen: removing cell materials with the aqueous solution soaking containing DNA enzymatic, after cleaning, then with containing α-gala The aqueous solution of glycosidase impregnates；It can also use 10--50mM aqueous slkali, under ultrasound condition, impregnate；Followed by ultrasonic with PBS Cleaning is until neutral；

5) semi-finished product are made to respective physical state on demand, then included dry, packaging, sterilizing and etc. be processed as finished product.

Further concrete operation method is:

1) it pre-processes, draws materials and clean: taking the small intestine or skin histology of breeding stock, sufficiently clean；Mechanical curettage or shearing method remove Non- connective tissue in small intestine, skin histology；By the submucous layer of small intestine isolated or skin corium；It rinses well, obtained target Tissue is exactly material feedstock；Then it places it in acetum and impregnates, soaking time 30--120 minutes, raw material was molten with acetic acid The ratio of liquid is 1:5--1:10(W/W)；

2) it sterilizes: with the mixed solution containing Peracetic acid and ethyl alcohol, in ultrasound and under room temperature, soak-out material raw material, into Row disinfection；Peroxyacetic acid concentration is 0.5-1.5%, concentration of alcohol 15-25%, and the ratio of material feedstock and mixed aqueous solution is 1:5--1:10, soaking time are 30--120 minutes；It is cleaned by ultrasonic again with purified water；

3) degreasing: using alcoholic solution, the soak-out material raw material under ultrasound, normal temperature condition, and material feedstock and alcoholic solution ratio are 1: 5--1:10, soak at room temperature time are 0.5--6h；It is cleaned by ultrasonic later with water；

4) remove cell: using the mixed solution of protease containing 0.05M and EDTA, under ultrasound, cryogenic conditions, soak-out material is former Material, is rinsed with PBS later；Repeatable 5 step 1--3 times；

5) it removes DNA and removes α-Gal antigen: the aqueous solution soaking material feedstock containing DNA enzymatic, after cleaning, then with containing alpha-galactoside The aqueous solution soaking material feedstock of enzyme is then used again and uses 10--30mM NaOH aqueous solution, under room temperature, ultrasound condition, impregnates material Expect raw material；Again with PBS ultrasonic cleaning until neutral；

6) semifinished material is made and presets required physical state, re-dry, packaging, sterilizing can obtain finished product.

Chinese and English term/noun first choice understands by following explanatory note, it is being separately described in detail unless；Other terms are pressed Those of ordinary skill in the art's level understands its meaning.

Chinese and English term explanation of nouns:

1) breeding stock: being for commodity livestock raised for meat；The main purpose of raising is for breeding cub, not as commodity Livestock raised for meat raises (such as pork pig feeding time is 5-6 months or so) in short term；Raising commodity livestock raised for meat main purpose be for putting on flesh, with For；Breeding stock, including pig, ox, sheep, horse, donkey, camel, dog；Signified breeding stock is all adult in this patent, numerous can educate, lead to Normal feeding time is more than at least a year；Such as boar includes sow and boar, sow here just refers to that reproducible adult is female Pig does not include young standby gilt；

2) through producing dam: belonging to a part of breeding stock, be the dam of at least raw a tire；Be for standby dam, Standby dam is primarily referred to as because age or monthly age arrive not yet, and there are no be pregnant young dam；

3) it eliminates dam: originally belonging to a part of breeding stock；In farm, manager is it is considered that when female livestock breed performance declines To a certain extent, as litter size is few, lactation amount is low, and weight of weaning litter is light, and comprehensively considers the utilization rate on dam column, eliminates sow The many factors such as price；Carry out discriminatory analysis from economics point, it is believed that it is significant to continue to raise dam value obtained It reduces, eliminating this kind of dam can be more cost-effective；Each farm has a certain difference the judgment criteria of superseded dam；Such as Usual sow can gradually be eliminated after 8 tires of giving a birth；Slightly has difference between the pig of different cultivars；

4) tissue renovation material, regeneration material, biological sticking patch, Biological Repair piece, biological support, degradable sticking patch, can inhale Sticking patch Bio-Mesh, Bio-Patch, Patch, Bioscaffold are received, this few Chinese and English noun or term, though have on surface Difference, but as its purpose and purposes be substantially；It is illustrated unless there are special, otherwise the intension of above-mentioned several nouns, It is substantially equivalent；

5) submucous layer of small intestine SIS(Small Intestinal Submucosa): small intestine includes jejunum and ileal segment, Remaining part, as the main component with collagen after removing mucous membrane of small intestine layer, muscle layer, placenta percreta, accounts for about 80% or more；

6) corium (Dermis): belong to a part of skin, between epidermis and hypodermic layer；From mesodermal differentiation, Divide papillaris pars and two layers of lamina reticularis, between the two without obvious boundary.1~2mm of the general thickness of corium；Mainly by fibroblast and its The collagenous fibres of generation, matrix are constituted, and have blood vessel, lymphatic vessel, nerve attachment.Matrix is by hyaluronic acid, chondroitin sulfate etc. The compound of glutinous polysaccharide and protein composition is constituted, and is filled in collagenous fibres and fibre bundle gap and iuntercellular, has hydrophily；

7) extracellular matrix ECM(Extracellular Matrix): be the acellular that is present in organized and organ at Point, required physical support not only is provided to cell tissue, is the normal physiological activity of various cells, suitable place is provided And microenvironment；And to the generation of the form of tissue, the chemotactic of cell and differentiation and important Physiology and biochemistry and biomethanics etc. Aspect plays important lever adjustment effect, thus the function of influence or organization of regulation control and organ.The function of cell 50% be by What the external microenvironment that extracellular matrix is built was determined.

Unless context has a clear explanation, the singular used in this specification and appended claims " one and "the" includes plural reference.Thus, for example, " this method " includes one or more methods and/or step, they belong to this paper institute It states type and/or is clearly will appreciate that after those skilled in the art have read this paper.Term " about " or " close " refer to Value range on statistical significance, range can be within an order of magnitude, usually specified number value or ranges 50% with It is interior, further refer within 20%, even more typically within 10%, or even is more typically within 5%.Term " about " or " close " The variation that can allow covered depends on the specific system of research, is that those skilled in the art one can easily realize that 's.

Combined with specific embodiments below, to further describe the principle of the present invention and scheme；It should be appreciated that these embodiments Thought of the invention is understood with facilitating solely for the purpose of illustration, but is not limited in this；Embodiment is not come in any way Limit the scope of the invention, in following embodiment, the various processes and method being not described in detail be it is used in the art it is known often The method of knowledge；It may also include being omitted to pass through what medical biotechnology Material Field used with gimmick and method.

Specific embodiment

Embodiment 1:(eliminates sow small intestine)

Selection is originated from the preparation for removing cell medical material of the submucous layer of small intestine of the superseded sow of health, the specific steps are as follows:

1) it pre-processes: in slaughterhouse, choosing the fresh small intestine for eliminating sow, the initial raw material after cleaning as sticking patch；Through It understands in depth, the common gestational age of the miscellaneous sow of superseded binary is more than 6 tires, and 180 kilograms of weight or more, be all the nonpregnant stage；It raises Feeding duration is usually more than 36 months；Mucous layer, muscle layer, placenta percreta, the lymph node that division removes chitterlings jejunum are scraped using physics Deng isolating submucosa, being placed in 0.5% acetum impregnates 30 minutes, and the ratio of chitterlings and acetum is 1:5, then makes It is rinsed 3 times with purified water, obtains biological sticking patch raw material, i.e. submucous layer of small intestine, following referred to as SIS materials；

2) it sterilizes: mixed aqueous solution of the use containing 1.0% Peracetic acid and 15% ethyl alcohol, SIS material and mixed aqueous solution Ratio is 1:10, under ultrasound condition, soaking at room temperature 100 minutes, is carried out disinfection.It is cleaned by ultrasonic 3 times using purified water later；

3) degreasing: the ethyl alcohol for the use of concentration being 90%, the ratio of SIS material and ethyl alcohol are 1:10, under ultrasound condition, soak at room temperature 2h.It is cleaned by ultrasonic 3 times using water for injection later；

4) cell is removed: using containing 0.02% trypsase and the mixed aqueous solution containing 0.02%EDTA, SIS material and trypsase/ The ratio of EDTA solution is 1:5, under ultrasound condition, impregnates 30 minutes, is impregnated 2 times with PBS solution later；Repetition is aforementioned to remove cell Step is secondary；

5) it removes DNA and removes α-Gal antigen: using the aqueous solution of the enzyme of DNA containing 5U/ml, the ratio of SIS material and DNA enzymatic solution For 1:5, under ultrasound condition, impregnated 20 minutes under the conditions of 37 DEG C；It is rinsed 3 times using PBS later；Use α containing 5U/ml-gala The ratio of the aqueous solution of glycosidase, SIS material and alpha-galactoside enzyme solutions is 1:5, under ultrasound condition, is soaked under the conditions of 30 DEG C Bubble 20 minutes；It is impregnated 2 times using PBS solution later；The NaOH aqueous solution that concentration is 10mM is reused, SIS material and NaOH are molten The ratio of liquid is 1:20, under ultrasound condition, soak at room temperature 50 minutes.Later using PBS ultrasonic cleaning until neutral；

6) it is formed, is lyophilized, sterilizing: the sheet-shaped material after cell being removed, four intersect in length and breadth, and overlapping is fixed on mold, freeze After drying, packaging finally carries out irradiation sterilization.

Embodiment 2:(Market pigs small intestine control group one)

The pretreatment of connective tissue, disinfection degreasing, remove cell, remove DNA and go α-Gal antigen, freeze-drying, sterilizing, completely With embodiment one；Difference only selects upper different from initial animal；It chooses in slaughterhouse, 100 kilograms of weight or so The fresh small intestine of the miscellaneous Market pigs of Ternary Pig ternary, cleans up；Understood it and raises number of days up to 160-180 days or so.

Embodiment 3:(eliminates sow --- corium)

Organize raw material pretreatment, disinfection, degreasing, remove cell, remove DNA and go α-Gal antigen, freeze-drying, sterilizing and etc. substantially With embodiment one；But it has any different in the first step and the 6th step；The distinctive points of the first step are type of feed difference, the present embodiment choosing Use the skin from slaughterhouse health superseded sow (live hog is again more than 180 kilograms) as initial raw material, position of drawing materials is pork pig Back middle part region；It is handled using the dermatome of profession, and removes destratum corneum and epidermis, and will be true using Mechanical Method 0.5mm is made in the thickness of cortex, in favor of subsequent every processing.

Second distinctive points is will to go sheet-shaped material two after cell in the 6th step, rather than use four as SIS Layer；Overlapping is fixed on mold, and after freeze-drying, packaging finally sterilizes.

Embodiment 4:(Market pigs corium control group two)

Organize raw material pretreatment, disinfection, degreasing, remove cell, remove DNA and go α-Gal antigen, freeze-drying, sterilizing and etc. substantially With embodiment one；But it has any different in the first step and the 6th step；The distinctive points of the first step are type of feed difference, the present embodiment choosing Use the skin from slaughterhouse health Market pigs (live hog weighs 100 kilograms or so) as initial raw material, position of drawing materials is pork pig Back middle part region；It is handled using the dermatome of profession, and removes destratum corneum and epidermis, and will using physical method 0.5mm is made in the thickness of skin corium, in favor of subsequent every processing.

Second distinctive points is will to go sheet-shaped material two after cell in the 6th step, rather than use four as SIS Layer；Overlapping is fixed on mold, and after freeze-drying, packaging finally sterilizes.

Embodiment 5: the mechanics properties testing of cell medical material is removed.

The detection for going cell medical material sample to carry out tensile strength that embodiment 1--4 is prepared.

Method: it is 10mm shape that sample is cut to width respectively in both directions；In relative humidity 40%- after cutting 60%, it is tested after being placed 2 hours in 22 ± 2 DEG C of environment of temperature.Fixture spacing is 25mm, and sample both ends are fixed on stretching Maximal force on the collet of testing machine, with 100mm/ minutes speed tensiles, when record is broken.

As a result it see the table below:

Those skilled in the art can make a variety of simple variations or adjustment or group to the present invention according to the above description It closes；Thus, under the premise of not violating claim objective of the invention, certain details in embodiment should not be constituted to this hair Bright restriction, the present invention will be using the ranges that the appended claims define as protection scope.

Claims (6)

1. a kind of medical acellular organism material, which is characterized in that the acellular organism material, with the tissue of breeding stock, organ Or combinations thereof object be primary raw material.

2. biomaterial according to claim 1, which is characterized in that the breeding stock is, sow, cow, ewe through producing, One of mare, female camel, jenny ass, female mule or combination.

3. biomaterial according to claim 1, which is characterized in that the breeding stock is superseded healthy sow, cow.

4. biomaterial according to claim 1, which is characterized in that the organ of the breeding stock be liver, pancreas, heart, One of kidney, lungs, tracheae or combination.

5. biomaterial according to claim 1, which is characterized in that the tissue of the breeding stock is submucous layer of small intestine, stomach One kind of submucosa, corium, pericardium, internal organs film, peritonaeum, pericystium or combinations thereof object.

6. a kind of preparation method of medical acellular organism material, which is characterized in that with the tissue of breeding stock, organ or combinations thereof object For primary raw material, cell technique is gone to be process.