CN101915693A - Human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining - Google Patents
Human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining Download PDFInfo
- Publication number
- CN101915693A CN101915693A CN2010102191679A CN201010219167A CN101915693A CN 101915693 A CN101915693 A CN 101915693A CN 2010102191679 A CN2010102191679 A CN 2010102191679A CN 201010219167 A CN201010219167 A CN 201010219167A CN 101915693 A CN101915693 A CN 101915693A
- Authority
- CN
- China
- Prior art keywords
- alcohol
- section
- hours
- nerve
- sample
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Abstract
The invention discloses a human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining. In the method, trigeminal ganglions, roots and branches of a fresh sample of a human embryo, especially an over 20-week induced-labor infant embryo not died with diseases are selected and subjected to HE conventional staining and observed under a light microscope and a transmission electron microscope, and finally the three-dimensional reconstruction is realized by using a computer TRI for win 95/NT, Version:3.01. By researching the form of the human embryonic trigeminus and advancing of motor roots, the invention shows the three-dimensional structure of the human embryonic trigeminus for the first time in human history, which not only enriches the research content of morphology of human embryonic trigeminus, but also provides a morphological basis for clinical prosopalgia pathophysiology and treatment. And thus, the human embryonic trigeminus based three-dimensional reconstruction method by using histotomy staining has a wide application scope.
Description
Technical field
The invention belongs to the neurotomy technical field.Specifically, the present invention relates to a kind ofly utilize tissue section strain to carry out the technical field of three-dimensional reconstruction based on people embryo trigeminal neuralgia.
Background technology
Trigeminal neuralgia (trigeminal nerve) is thick Combination cranial nerve, contains two kinds of fibers of general somatesthesia and special internal organ motion.Special visceral motor fiber arises from the nuclei motorii nervi trigemini in pons stage casing, its aixs cylinder is formed motor root of trigeminal nerve, basilar part of pons (from the pons facies ventralis) goes out brain with the medipeduncle place of dividing a word with a hyphen at the end of a line, be positioned at the preceding inboard of sensory root, enter mandibular nerve at last, go out cranium through oval foramen, branch is distributed in masseter etc. with mandibular nerve.The cell space of general somatesthesia fiber concentrates in the gasserian ganglion, and wherein central process is concentrated and constituted thick radix sensoria nervi trigemini, goes into brain by basilar part of pons and medipeduncle intersection, terminates in all sensory nucleus of trigeminal neuralgia; The prominent trigeminal neuralgia three big branches that form around the trigeminal neuralgia ganglion cell, i.e. the 1st ophthalmic nerve, the 2nd maxillary nerve, the 3rd is mandibular nerve.From 3 continuous branches of big branch be distributed in skin of face, eye and the socket of the eye, the mucous membrane of oral cavity, nasal cavity, paranasal sinus, tooth, meninxes etc., polyesthesia such as touch at conduction pain, temperature.
(trigeminal neuralgia is modal a kind of in the cranial neuralgia TN) to trigeminal neuralgia, also is one of disease common in the peripheral nervous disease.According to statistics, this sick morbidity rate is 1,82/,100,000, morbidity is a feature with the lightening violent facial pain that takes place suddenly in one of facial trigeminal neuralgia or several distributive province, the patient often be described as tear sample, electric shock sample, lightning sample, acupuncture sample, sample or burn sample severe pain lancinates.Pain is the most obvious with cheek, the upper jaw, lower jaw or tongue.Because the definite pathogenesis of trigeminal neuralgia and the cause of disease it be unclear that, thus desirable methods of treatment lacked clinically, common with surgical operation therapy.The selectivity sensory rhizotomy of trigeminal nerve is one of surgical intervention trigeminal neuralgia method commonly used at present.But can damage motor root sometimes in the art, show as the homonymy masticatory paralysis, dental articulation is not tight, and some patient Yi Fasheng temporomandibular joint dislocation also has some patients difficulty in opening mouth can occur, and lower jaw is partial to Ipsilateral etc. when dehiscing.Need to differentiate the relation that the interior sensation of gasserian ganglion is propped up and motion is propped up in the art.Therefore, research, the especially motor root of trigeminal nerve traveling in gasserian ganglion of Roots of trigeminal nerve morphology aspect has certain theory and practice significance to the prosopalgic cause of disease and treatment.
The traveling of adult's form of Roots of trigeminal nerve and motor root of trigeminal nerve has been received the concern of many Chinese scholars, and this has been carried out a series of research.People such as Young find in limited time that in the motion of research adult trigeminal neuralgia motor root contains the Remak's nerve fiber of some, these Remak's nerve fibers are unknowable in the effect of Roots of trigeminal nerve, but have circumstantial evidence to show, these Remak's nerve fibers have potential sensory function, and this may be relevant with radix sensoria nervi trigemini selective rhizotomy postoperative persistence of sensation and pain sensation alleviation failure.Ezure etc. discover big than sensory root of the area of aixs cylinder in the Roots of trigeminal nerve and girth, and the aixs cylinder distribution range is wider than sensory root, no thick a fiber in the sensory root.Sun Eryu etc. utilize adult's corpse head as sample, and the utilization surgery microscope has been done careful anatomical research to the trigeminal neuralgia intracranial segment, and motor root and sensation location of root relation have been discussed, and think to have ramus anastomoticus between motor root and the sensory root.But, do not see people embryo Roots of trigeminal nerve and motor root traveling correlative study report so far both at home and abroad as yet from the viewpoint of auxology.
Summary of the invention
At embryo's Roots of trigeminal nerve and the motor root traveling correlative study report of also having no talent at present.The present invention is based on people embryo trigeminal neuralgia and utilize tissue section strain to realize three-dimensional reconstruction, obtained effect preferably.
The invention provides and a kind ofly utilize tissue section strain to carry out three-dimensional rebuilding method based on people embryo trigeminal neuralgia.By selecting people embryo fresh specimens, be the induced labor foetus of non-disease death especially, the gasserian ganglion of the gestational age people embryo fresh specimens that 20 weeks are above, root and nervous ramification, through the HE normal dyeing, utilize light border transmission electron microscope observing, adopt the computer realization three-dimensional reconstruction.
The present invention is by Roots of trigeminal nerve, joint and the HE of branch normal dyeing to sample, utilize light microscopic, electron microscopic observation, three-dimensional reconstruction, the research of method by the traveling of trifacial form of people embryo and motor root is carried out, be for the first time the trifacial 3-D solid structure of people embryo to be shown on the human history, both enrich the content of human trigeminal neuralgia morphology research, provided morphologic basis for clinical prosopalgic pathologic, physiologic and treatment simultaneously.
The present invention provides specifically that a kind of to utilize tissue section strain to carry out the three-dimensional rebuilding method step based on people embryo trigeminal neuralgia as follows:
1. sample is fixed
In the 1-4h, with after the flushing of physiological saline heart, fixing through aortic perfusion with 10% formalin solution 1000-1500mL more earlier after obtaining sample, keeps in Dark Place in dry place.
2. visual inspection
After treating that sample is completely fixed, get fetal head, open cranium, expose akrencephalon, win a side cerebral hemisphere and a diencephalon, remove the part midbrain, determine that left side Roots of trigeminal nerve, joint and three big branches carry out big and small dissection and observation, carry out material object and take pictures at the surgery microscopically.
3. electron microscopic observation
In the complete taking-up of surgery microscopically right side Roots of trigeminal nerve, gasserian ganglion and three big branches, after finishing was organized, naked eyes can be distinguished motor root of trigeminal nerve, peel off motor root of trigeminal nerve with the sample that obtains; Organize respectively with what get that mark places the container that 2.5% glutaraldehyde solution is housed well, the set time is 72 hours; Tissue that fixation mark is good takes out, and moves into the rinsing 3 times in PH7.4 of 1/15M phosphate buffer, each each 15 minutes; After fixing 1 hour after being organized in the 1% hungry sour immobile liquid after the rinsing, by as above entering dehydration after the step rinsing; After entering 50% alcohol, 70% alcohol, 80% alcohol, 90% alcohol, 100% alcohol and respectively dewatering 15 minutes, be placed in 100% alcohol and dewater; In v/v, adopt acetone and epoxy resin to soak in the liquid 1 hour according to 1: 1, acetone and epoxy resin in liquid soaked into 3 hours at 1: 3, soak into 12 hours in the pure epoxy resin after; Adopt the pure epoxy resin embedding, 37 ℃ 12 hours, 45 ℃ 12 hours, 60 ℃ 24 hours; Employing ultramicrotome section selects wherein that 1-2 opens section, observes under optical microscope after the Toluidine blue staining, according to the foundation that the Toluidine blue staining result provides, selects the more position of neuron to carry out ultra-thin section again; Select for use the two colouring methods of uranium acetate and lead citrate that section is dyeed; Section is placed transmission electron microscope observing, gather picture.
4.HE dyeing
Each sample all with the filter paper parcel, with after soaking 12 hours in the crocus cloth parcel water, was gone into 75% dehydration of alcohol 4 hours outward, 85% dehydration of alcohol 4 hours, 90% alcohol spends the night, and is placed on 95% dehydration of alcohol 4 hours, and 100% dehydration of alcohol spends the night with 100% alcohol after 4 hours again; Went among the dimethylbenzene I 30 minutes, and went among the dimethylbenzene II till transparent; Went among the electric oven liquid wax I saturating wax 1 hour, saturating wax is 30 minutes among the liquid wax II; Take out sample and put into embedding frame wax oil and treat that it puts into tap water after fixing and cool off, treat that wax stone is shaped after, wax stone is fixed mark, place refrigerator and cooled to hide, use during section; Select microtome for use, make serial section by coronal section, slice thickness is 6 μ m; The section of mounting is placed on the staining rack, put into 60 ℃ of baking boxs 30 minutes.Go in dimethylbenzene I, II, the III liquid each after the taking-up 2 minutes, and went into 100% alcohol I, II, 95% alcohol, each 1-2 of 85% alcohol minute, use tap water, distilled water flushing a moment respectively.Su Dasu contaminated 5 minutes, and the tap water developing sheet is carved into 1% hydrochloride alcohol differentiation 3-5 second, and the tap water flushing for a moment.Ammoniacal liquor a moment, tap water flushing a moment to go into 0.5% Yihong solution and contaminated 5 minutes, 75% alcohol, 85% alcohol, 95% alcohol I are moments later gone in washing, II, 100% alcohol I, dehydration in each 1-2 of II minute reenters among dimethylbenzene I, the II transparent each 1-2 minute.Neutral gum sealing section preparation is again with every biopsy marker ordering; Section is placed observation under the light microscopic, gather picture.
5. three-dimensional reconstruction
Utilize the HE coloration result, under light microscopic,, form the former figure of every section with the automated imaging technology in 40 times of visual field images acquired; Adopt similar Adobe photoshop instrument to the different colors of usefulness such as cell, nerve root, each branch and motor root of each section boundary line of drawing; Each that adopts that similar Adobe photoshop instrument finishes is opened section by from the bottom up rank order, with each section according to cell, three big branches, motion location of root to good, draw a triangle on the tissue next door then, the position of this triangle from first to a last section must be consistent, it is preserved with the psd form as a standard axle; With each image size 2446 * 2669 that the psd form is preserved, pixel 72,536KB changes 562 * 629 into, pixel 100,52.2KB, amended image is preserved with the jpg form.Every image that the jpg form is preserved is imported computing machine in order, select the slice thickness of rebuilding.
The used organization material of the present invention adopts the coronal section serial section, bed thickness 6 μ m since at interval three choose picture, so the bed thickness of actual reconstruction is 18 μ m, finishes the three-dimensional image reconstruction with TRI for Win 95/NT Version:3.10 software then.
Can clearly show the distribution of people embryo Roots of trigeminal nerve, joint and branch thereof by the three-dimensional image of the present invention's reconstruction, the piece cutting structure of people embryo Roots of trigeminal nerve, joint and the HE of branch dyeing is rebuild out, demonstrated the corresponding relation between the three better.People embryo's motor root of trigeminal nerve and sensory root companion row as can be seen from the picture of three-dimensional reconstruction, be positioned at the below of sensory root, and the dark face in 1/3 place is kept straight on below gradually in the trigeminal neuralgia semilunar ganglion, is parallel to the dark face of preceding inner edge of mandibular nerve afterwards, is slightly covered.The three big branches that the gasserian ganglion cell process is formed are respectively ophthalmic nerve, maxillary nerve and mandibular nerve under outside interior.
By implementing the concrete content of the present invention, can reach following beneficial effect.
1, the three-dimensional image of the present invention's reconstruction can clearly show the distribution of people embryo Roots of trigeminal nerve, joint and branch thereof, 1/3 place under the concentrated inboard of motor root of trigeminal nerve, the neuromere by radix sensoria nervi trigemini, be a continuous structure from the root to the mandibular nerve, incorporate the interior below of mandibular nerve at last into.Thereby show that the method that the present invention utilizes the sample of histotomy to carry out three-dimensional reconstruction is feasible.
2, traditional three-dimensional reconstruction has method not of the same race, the related personnel of non-computer specialty is difficult to independently finish three-dimensional reconstruction, the inventive method is simple and practical, general laboratory technician, amateur computernik can finish complicated histotomy three-dimensional reconstruction, and method is specifically feasible, can independently finish.
3, the present invention rebuilds by three-dimensional image, can clearly see Roots of trigeminal nerve, joint and branch's 3-D solid structure, especially can see also that from the dorsal part of sample the trigeminal neuralgia motor fiber is at Roots of trigeminal nerve, position traveling within joint and the mandibular nerve, this provides the morphologic basis of aspects such as relevant dissection, histology to prosopalgic surgical intervention, and this operation is had certain theory and practice meaning.
Description of drawings
Fig. 1 demonstration is the location drawing of people embryo Roots of trigeminal nerve, joint and three branches, and among the figure, V is a Roots of trigeminal nerve; V1 is an ophthalmic nerve; V2 is a maxillary nerve; V3 is a mandibular nerve; GN is a gasserian ganglion; II is an optic nerve; III is an oculomotor nerve; P is a pons; C is a cerebellum; M is a midbrain; T is an akrencephalon.
Fig. 2 is shown as Roots of trigeminal nerve, joint and the branch aspect graph under light microscopic, and under 4 * 10 times of conditions of HE dyeing, among the figure, Vr is a Roots of trigeminal nerve; V1 is an ophthalmic nerve; V2 is a maxillary nerve; V3 is a mandibular nerve; GN is a gasserian ganglion; Mr is the kinesitherapy nerve root.
Embodiment
Below with reference to accompanying drawing structure composition of the present invention and mode of operation thereof and principle are described further, still, the present invention is not limited to following embodiment.In addition, in following explanation, if no special instructions, % all refers to the V/V percent by volume.
Equipment and material have among the present invention:
Collect non-disease terminal pregnancy and the healthy fetus of induced labor, male or female, gestational age are more than 20 weeks, and gestational age is obtained by the health check-up of ultrasound diagnosis in conjunction with sample according to pregnant woman's last menstrual period.All samples of the present invention are fetched and delivered the laboratory at once all from the municipal district hospital, Urumchi after the fetus induced labor, earlier with behind the normal saline flushing, fix through aortic perfusion with 10% formalin solution 1000-1500mL again.Long for fear of the sample death time, tissue degeneratiaon influences test findings, and the present invention is controlled within 4 hours to time of perfusion fixation after with the sample induced labor.
Surgery microscope: No.1 Hospital Attached to Xinjiang Medical Univ. provides, micro-surgical instruments: Cao worker's board, the kind boat of Shandong Zibo engineering in medicine research institute makes, the routine operation apparatus: human dissection teaching and research room of Xinjiang Medicine University provides, fluorescent microscope: Xiamen Motic BA400 type, autopsy table: human dissection teaching and research room of Xinjiang Medicine University provides, digital camera: Japanese Nikon 4500 digital cameras (microspur 2cm), paraffin slicing machine: German Lycra 2135 types, transmission electron microscope: Japanese JEM-1230 type, ultramicrotome: Sweden LKB-2188 type, full microscope: Xiamen Motic BA600 type, refrigerator: Qingdao Haier, electronic analytical balance: plum Teller-Tuo benefit Instr Ltd., MLS-3750 type laboratory autoclave: Sanyo Electric International Trading Company Ltd, three-dimensional reconstruction computing machine: the Japanese medical university second anatomy classroom
The reagent that relates among the present invention and the preparation of working fluid:
Chemical reagent: Dil:FMP FM003282, Japanese medical university provides, the PBS pulvis: Fuzhou Maixin biotechnology Development Co., Ltd, formalin analyze pure: Xi'an chemical reagent factory, absolute ethyl alcohol: Xi'an chemical reagent factory, dimethylbenzene, Yihong, haematoxylin, sodium chloride, potassium chloride, sodium hydrogen phosphate, potassium dihydrogen phosphate etc. are pure for homemade analysis, distilled water, 2.5% glutaraldehyde, 10% formalin solution etc. provide for human dissection teaching and research room of Xinjiang Medicine University and Electron Microscopy Room.
All reagent selected for use among the present invention and instrument all are well known selecting for use, but do not limit enforcement of the present invention, and other reagent more well known in the art and equipment are all applicable to the enforcement of the following embodiment of the present invention.
Embodiment one: utilize tissue section strain to carry out three-dimensional rebuilding method based on people embryo trigeminal neuralgia
1. sample is fixed
After obtaining sample in the 1-4h, clamp fetal cord with pliers, open the front wall, expose pericardium, cut pericardium, fully expose heart, No. 7 infusion niidls are inserted root of ascending aorta, cut a little otch with scissors in right atrium, after injecting the flushing of the capable heart of 0.9% physiological saline 500ml continuously with the 50ml syringe, inject 10% formalin solution 1500ml again, allow immobile liquid along the capable perfusion fixation of body circulation approach, observe the fixed effect of sample, see the mouth of immobile liquid from sample arranged, noses etc. are located to flow out sample abdominal distention, this good fixing effect of limbs hardening indicating, the sample that perfusion is good continues to be soaked in that to carry out back in 10% formalin solution of fresh configuration fixing, and dry locating kept in Dark Place.
2. visual inspection
After treating that 4 routine samples are completely fixed, get fetal head, open cranium, expose akrencephalon, win a side cerebral hemisphere and a diencephalon, remove the part midbrain at the surgery microscopically, determine that left side Roots of trigeminal nerve, joint and three big branches carry out big and small dissection and observation, carry out material object and take pictures, referring to accompanying drawing 1,2.
3. electron microscopic observation
(1) draw materials: above-mentioned 4 routine samples are in the complete taking-up of surgery microscopically right side Roots of trigeminal nerve, gasserian ganglion and three big branches, behind the finishing tissue, naked eyes can be distinguished motor root of trigeminal nerve, peel off motor root of trigeminal nerve, and traveling is in the kinesitherapy nerve root of trigeminal neuralgia sections.
(2) preceding fixing: that will get organizes respectively that mark places the container that 2.5% glutaraldehyde solution is housed well, and the set time is 72 hours.
(3) rinsing: tissue that fixation mark is good takes out, and moves into the rinsing 3 times in PH7.4 of 1/15M phosphate buffer, each each 15 minutes.
(4) back is fixing: fix 1 hour after being organized in the 1% hungry sour immobile liquid after the rinsing.
(5) rinsing: operation steps is with (3).
(6) dehydration: go into 50% alcohol, 70% alcohol, 80% alcohol, 90% alcohol, 100% alcohol I and respectively dewatered 15 minutes, be placed among the 100% alcohol II.
(7) soak into: according to volume ratio, acetone and epoxy resin in liquid soaked into 1 hour at 1: 1, and acetone and epoxy resin in liquid soaked into 3 hours at 1: 3, soaked in the pure epoxy resin 12 hours.
(8) embedding: the pure epoxy resin embedding, 37 ℃ 12 hours, 45 ℃ 12 hours, 60 ℃ 24 hours.
(9) section: with the section of Sweden LKB-2188 type ultramicrotome, select wherein that 1-2 opens section, under optical microscope, observe after the Toluidine blue staining,, select the more position of neuron to carry out ultra-thin section again according to the foundation that the Toluidine blue staining result provides.
(10) dyeing: select for use the two colouring methods of uranium acetate and lead citrate that section is dyeed.
(11) observe: section is placed Japanese JEM-1230 type transmission electron microscope observing, gather picture.
4.HE dyeing three-dimensional reconstruction
(1) draws materials: 4 routine samples, after waiting to be completely fixed, get fetal head, after the PBS rinsing is clean, remove scalp along zygomatic arch upper limb 1cm plane annular, take off braincap, cut off the endocranium of braincap portion again along same plane, the cross-section brain stem of flat superior colliculus upper limb, excision bilateral akrencephalon and diencephalon, the careful endocranium that cuts off tentorium cerebelli and gasserian ganglion top, will be together with the basis cranii of brain stem, Roots of trigeminal nerve, joint, the position of propping up is without any change, place under the surgery microscope, 2 routine samples are determined the left side Roots of trigeminal nerve, gasserian ganglion and three big branch ophthalmic nerves, maxillary nerve, the position of mandibular nerve, other 2 routine samples are determined the right side Roots of trigeminal nerve, gasserian ganglion and three big branch ophthalmic nerves, maxillary nerve, the position of mandibular nerve, cut off ophthalmic nerve respectively at the superior orbital fissure place, the circular hole place cuts off maxillary nerve, the oval foramen place cuts off mandibular nerve, basilar part of pons cuts off Roots of trigeminal nerve, the careful complete Roots of trigeminal nerve of peeling off, gasserian ganglion and three big branches, behind the finishing tissue, be positioned in 10% formalin solution of fresh configuration.
(2) embedding: each sample with after soaking 12 hours in the crocus cloth parcel water, was gone into 75% dehydration of alcohol 4 hours outward all with the filter paper parcel, 85% dehydration of alcohol 4 hours, 90% alcohol spends the night, and is placed on 95% dehydration of alcohol 4 hours, 100% alcohol I dehydration 4 hours, 100% alcohol II spends the night; Went among the dimethylbenzene I 30 minutes, and went among the dimethylbenzene II till transparent; Went among the electric oven liquid wax I saturating wax 1 hour, saturating wax is 30 minutes among the liquid wax II; Take out sample and put into embedding frame wax oil and treat that it puts into tap water after fixing and cool off, treat that wax stone is shaped after, wax stone is fixed mark, place refrigerator and cooled to hide, use during section.
(3) section: with section center Germany Lycra 2135 type microtomes, make serial section by coronal section, slice thickness is 6 μ m.Every routine Roots of trigeminal nerve, neuromere sample all have the serial section about 130 in the 4 routine samples.
(4) dyeing: the section that will mount places on the staining rack, puts into 60 ℃ of baking boxs 30 minutes.Go in dimethylbenzene I, II, the III liquid each after the taking-up 2 minutes, and went into 100% alcohol I, II, 95% alcohol, each 1-2 of 85% alcohol minute, use tap water, distilled water flushing a moment respectively.Haematoxylin was contaminated 5 minutes, and the tap water developing sheet is carved into 1% hydrochloride alcohol differentiation 3-5 second, and the tap water flushing for a moment.Ammoniacal liquor a moment, tap water flushing a moment to go into 0.5% Yihong solution and contaminated 5 minutes, 75% alcohol, 85% alcohol, 95% alcohol I are moments later gone in washing, II, 100% alcohol I, dehydration in each 1-2 of II minute reenters among dimethylbenzene I, the II transparent each 1-2 minute.Neutral gum sealing section preparation is again with every biopsy marker ordering.
(5) observe: section is placed under the light microscopic observe, gather picture.
5. three-dimensional reconstruction
(1) HE coloration result in 40 times of visual field images acquired, uses the automated imaging technology to form the former figure of every section, totally 136 under light microscopic.
(2) with Adobe photoshop 7.0 to the different colors of usefulness such as cell, nerve root, each branch and motor root of each section boundary line of drawing.
(3) each that finish with Adobe photoshop 7.0 is opened section by from the bottom up rank order, with the position of each section to good, as cell, three big branches, motor root etc., draw a triangle on the tissue next door then, the position of this triangle from first to a last section must be consistent, it is preserved with the psd form as a standard axle.
(4) each pictures life size 2446 * 2669, pixel 72,536KB changes 562 * 629 into, pixel 100,52.2KB must preserve with the jpg form.
(5) finish three-dimensional image with TRI for Win 95/NT Version:3.10.
Embodiment two: sample is fixed
After obtaining sample in the 1-4h, clamp fetal cord with pliers, open the front wall, expose pericardium, cut pericardium, fully expose heart, No. 7 infusion niidls are inserted root of ascending aorta, cut a little otch with scissors in right atrium, after injecting the flushing of the capable heart of 0.9% physiological saline 500ml continuously with the 50ml syringe, inject 10% formalin solution 1500ml again, allow immobile liquid along the capable perfusion fixation of body circulation approach, observe the fixed effect of sample, see the mouth of immobile liquid from sample arranged, noses etc. are located to flow out sample abdominal distention, this good fixing effect of limbs hardening indicating, the sample that perfusion is good continues to be soaked in that to carry out back in 10% formalin solution of fresh configuration fixing, and dry locating kept in Dark Place.
Embodiment three: light microscopic, transmission electron microscope observing
(1) draw materials: the foregoing description sample is in the complete taking-up of surgery microscopically right side Roots of trigeminal nerve, gasserian ganglion and three big branches, behind the finishing tissue, naked eyes can be distinguished motor root of trigeminal nerve, peel off motor root of trigeminal nerve, traveling is in the kinesitherapy nerve root of trigeminal neuralgia sections, referring to accompanying drawing 1,2.
(2) preceding fixing: that will get organizes respectively that mark places the container that 2.5% glutaraldehyde solution is housed well, and the set time is 72 hours.
(3) rinsing: tissue that fixation mark is good takes out, and moves into the rinsing 3 times in PH7.4 of 1/15M phosphate buffer, each each 15 minutes.
(4) back is fixing: fix 1 hour after being organized in the 1% hungry sour immobile liquid after the rinsing.
(5) rinsing: operation steps is with (3).
(6) dehydration: go into 50% alcohol, 70% alcohol, 80% alcohol, 90% alcohol, 100% alcohol I and respectively dewatered 15 minutes, be placed among the 100% alcohol II.
(7) soak into: according to volume ratio, acetone and epoxy resin in liquid soaked into 1 hour at 1: 1, and acetone and epoxy resin in liquid soaked into 3 hours at 1: 3, soaked in the pure epoxy resin 12 hours.
(8) embedding: the pure epoxy resin embedding, 37 ℃ 12 hours, 45 ℃ 12 hours, 60 ℃ 24 hours.
(9) section: with the section of Sweden LKB-2188 type ultramicrotome, select wherein that 1-2 opens section, under optical microscope, observe after the Toluidine blue staining,, select the more position of neuron to carry out ultra-thin section again according to the foundation that the Toluidine blue staining result provides.
(10) dyeing: select for use the two colouring methods of uranium acetate and lead citrate that section is dyeed.
(11) observe: section is placed Japanese JEM-1230 type transmission electron microscope observing, gather picture.
Embodiment four: HE dyeing
(1) draws materials: 4 routine samples, after waiting to be completely fixed, get fetal head, after the PBS rinsing is clean, remove scalp along zygomatic arch upper limb 1cm plane annular, take off braincap, cut off the endocranium of braincap portion again along same plane, the cross-section brain stem of flat superior colliculus upper limb, excision bilateral akrencephalon and diencephalon, the careful endocranium that cuts off tentorium cerebelli and gasserian ganglion top, will be together with the basis cranii of brain stem, Roots of trigeminal nerve, joint, the position of propping up is without any change, place under the surgery microscope, 2 routine samples are determined the left side Roots of trigeminal nerve, gasserian ganglion and three big branch ophthalmic nerves, maxillary nerve, the position of mandibular nerve, other 2 routine samples are determined the right side Roots of trigeminal nerve, gasserian ganglion and three big branch ophthalmic nerves, maxillary nerve, the position of mandibular nerve, cut off ophthalmic nerve respectively at the superior orbital fissure place, the circular hole place cuts off maxillary nerve, the oval foramen place cuts off mandibular nerve, basilar part of pons cuts off Roots of trigeminal nerve, the careful complete Roots of trigeminal nerve of peeling off, gasserian ganglion and three big branches, behind the finishing tissue, be positioned in 10% formalin solution of fresh configuration.
(2) embedding: each sample with after soaking 12 hours in the crocus cloth parcel water, was gone into 75% dehydration of alcohol 4 hours outward all with the filter paper parcel, 85% dehydration of alcohol 4 hours, 90% alcohol spends the night, and is placed on 95% dehydration of alcohol 4 hours, 100% alcohol I dehydration 4 hours, 100% alcohol II spends the night; Went among the dimethylbenzene I 30 minutes, and went among the dimethylbenzene II till transparent; Went among the electric oven liquid wax I saturating wax 1 hour, saturating wax is 30 minutes among the liquid wax II; Take out sample and put into embedding frame wax oil and treat that it puts into tap water after fixing and cool off, treat that wax stone is shaped after, wax stone is fixed mark, place refrigerator and cooled to hide, use during section.
(3) section: with section center Germany Lycra 2135 type microtomes, make serial section by coronal section, slice thickness is 6 μ m.Every routine Roots of trigeminal nerve, neuromere sample all have the serial section about 130 in the 4 routine samples.
(4) dyeing: the section that will mount places on the staining rack, puts into 60 ℃ of baking boxs 30 minutes.Go in dimethylbenzene I, II, the III liquid each after the taking-up 2 minutes, went into 100% alcohol I, 100% alcohol II, 95% alcohol, each 1-2 of 85% alcohol minute, use tap water, distilled water flushing a moment respectively.Haematoxylin was contaminated 5 minutes, and the tap water developing sheet is carved into 1% hydrochloride alcohol differentiation 3-5 second, and the tap water flushing for a moment.Ammoniacal liquor a moment, tap water flushing a moment, go into 0.5% Yihong solution and contaminated 5 minutes, 75% alcohol, 85% alcohol, 95% alcohol I are moments later gone in washing, 95% alcohol II, 100% alcohol I, dehydration in each 1-2 of II minute reenters among dimethylbenzene I, the II transparent each 1-2 minute.Neutral gum sealing section preparation is again with every biopsy marker ordering.
(5) observe: section is placed under the light microscopic observe, gather picture.
Embodiment five: three-dimensional reconstruction
(1) HE coloration result in 40 times of visual field images acquired, uses the automated imaging technology to form the former figure of every section, totally 136 under light microscopic.
(2) with Adobe photoshop 7.0 to cell, nerve root, each branch and the motor root etc. of each section with different boundary lines, color picture place.
(3) each that finish with Adobe photoshop 7.0 is opened section by from the bottom up rank order, with the position of each section to good, as cell, three big branches, motor root etc., draw a triangle on the tissue next door then, the position of this triangle from first to a last section must be consistent, it is preserved with the psd form as a standard axle.
(4) each pictures life size 2446 * 2669, pixel 72,536KB changes 562 * 629 into, pixel 100,52.2KB must preserve with the jpg form.
(5) finish three-dimensional image with TRI for Win 95/NT Version:3.10.
Claims (2)
1. one kind is utilized tissue section strain to carry out three-dimensional rebuilding method based on people embryo trigeminal neuralgia, by Roots of trigeminal nerve, joint and branch's dyeing to sample, utilize light microscopic, electron microscopic observation, three-dimensional reconstruction, it is characterized in that, according to the percent by volume meter, described to utilize tissue section strain to carry out the three-dimensional reconstruction concrete steps based on people embryo trigeminal neuralgia as follows:
(1) sample is fixed: in the 1-4h, with after the flushing of physiological saline heart, fixing through aortic perfusion with 10% formalin solution 1000-1500mL more earlier after obtaining sample, keeps in Dark Place in dry place;
(2) visual inspection: after treating that sample is completely fixed, get fetal head, open cranium, expose akrencephalon, win a side cerebral hemisphere and a diencephalon, remove the part midbrain at the surgery microscopically, determine that left side Roots of trigeminal nerve, joint and three big branches carry out big and small dissection and observation, carry out material object and take pictures;
(3) electron microscopic observation: in the complete taking-up of surgery microscopically right side Roots of trigeminal nerve, gasserian ganglion and three big branches, after finishing was organized, naked eyes can be distinguished motor root of trigeminal nerve, peel off motor root of trigeminal nerve with the sample that obtains; Organize respectively with what get that mark places the container that 2.5% glutaraldehyde solution is housed well, the set time is 72 hours; Tissue that fixation mark is good takes out, and moves into the rinsing 3 times in PH7.4 of 1/15M phosphate buffer, each each 15 minutes; After fixing 1 hour after being organized in the 1% hungry sour immobile liquid after the rinsing, by as above entering dehydration after the step rinsing; After entering 50% alcohol, 70% alcohol, 80% alcohol, 90% alcohol, 100% alcohol and respectively dewatering 15 minutes, be placed in 100% alcohol and dewater; In v/v, adopt acetone and epoxy resin to soak in the liquid 1 hour according to 1: 1, acetone and epoxy resin in liquid soaked into 3 hours at 1: 3, soak into 12 hours in the pure epoxy resin after; Adopt the pure epoxy resin embedding, 37 ℃ 12 hours, 45 ℃ 12 hours, 60 ℃ 24 hours; Employing ultramicrotome section selects wherein that 1-2 opens section, observes under optical microscope after the Toluidine blue staining, according to the foundation that the Toluidine blue staining result provides, selects the more position of neuron to carry out ultra-thin section again; Select for use the two colouring methods of uranium acetate and lead citrate that section is dyeed; Section is placed transmission electron microscope observing, gather picture;
(4) HE dyeing: each sample all with the filter paper parcel, with after soaking 12 hours in the crocus cloth parcel water, was gone into 75% dehydration of alcohol 4 hours outward, 85% dehydration of alcohol 4 hours, 90% alcohol spends the night, and is placed on 95% dehydration of alcohol 4 hours, and 100% dehydration of alcohol spends the night with 100% alcohol after 4 hours again; Went among the dimethylbenzene I 30 minutes, and went among the dimethylbenzene II till transparent; Went among the electric oven liquid wax I saturating wax 1 hour, saturating wax is 30 minutes among the liquid wax II; Take out sample and put into embedding frame wax oil and treat that it puts into tap water after fixing and cool off, treat that wax stone is shaped after, wax stone is fixed mark, place refrigerator and cooled to hide, use during section; Select microtome for use, make serial section by coronal section, slice thickness is 6 μ m; The section of mounting is placed on the staining rack, put into 60 ℃ of baking boxs 30 minutes.Go in dimethylbenzene I, II, the III liquid each after the taking-up 2 minutes, went into 100% alcohol I, alcohol II, 95% alcohol, each 1-2 of 85% alcohol minute, use tap water, distilled water flushing a moment respectively.Haematoxylin was contaminated 5 minutes, and the tap water developing sheet is carved into 1% hydrochloride alcohol differentiation 3-5 second, and the tap water flushing for a moment.Ammoniacal liquor for a moment, the tap water flushing for a moment, go into 0.5% Yihong solution and contaminated 5 minutes, 75% alcohol, 85% alcohol, 95% alcohol I, 95% alcohol II, 100% alcohol I, 100% each 1-2 of alcohol II minute dehydration are moments later gone in washing, reenter among dimethylbenzene I, the II transparent each 1-2 minute; Neutral gum sealing section preparation is again with every biopsy marker ordering; Section is placed observation under the light microscopic, gather picture;
(5) three-dimensional reconstruction: utilize the HE coloration result, under light microscopic,, form the whole former figure of every section with the automated imaging technology in 40 times of visual field images acquired; Adopt similar Adobe photoshop instrument to the different colors of usefulness such as cell, nerve root, each branch and motor root of each section boundary line of drawing; Each that adopts that similar Adobe photoshop instrument finishes is opened section by from the bottom up rank order, with each section according to cell, three big branches, motion location of root to good, draw a triangle on the tissue next door then, the position of this triangle from first to a last section must be consistent, it is preserved with the psd form as a standard axle; With each image size 2446 * 2669 that the psd form is preserved, pixel 72,536KE changes 562 * 629 into, pixel 100,52.2KB, amended image is preserved with the jpg form; Every image that the jpg form is preserved is imported computing machine in order, select the slice thickness of rebuilding.
2. utilize tissue section strain to carry out three-dimensional rebuilding method as claim 1 based on people embryo trigeminal neuralgia, it is characterized in that, described organization material adopts the coronal section serial section, bed thickness 6 μ m, choose picture for three at interval, the bed thickness of rebuilding is 18 μ m, finishes three-dimensional image with TRI for Win 95/NT Version:3.10 software then and rebuilds.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010102191679A CN101915693B (en) | 2010-07-07 | 2010-07-07 | Human embryonic trigeminus based three-dimensional reconstruction method by using slide stainer |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010102191679A CN101915693B (en) | 2010-07-07 | 2010-07-07 | Human embryonic trigeminus based three-dimensional reconstruction method by using slide stainer |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101915693A true CN101915693A (en) | 2010-12-15 |
| CN101915693B CN101915693B (en) | 2012-07-25 |
Family
ID=43323255
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2010102191679A Expired - Fee Related CN101915693B (en) | 2010-07-07 | 2010-07-07 | Human embryonic trigeminus based three-dimensional reconstruction method by using slide stainer |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101915693B (en) |
Cited By (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102243154A (en) * | 2011-04-02 | 2011-11-16 | 中山大学附属第一医院 | Method for dyeing peripheral nerves slice |
| CN102519774A (en) * | 2011-11-16 | 2012-06-27 | 中国水产科学研究院东海水产研究所 | Dyeing method for peripheral nerves of fish larvae and juveniles |
| CN102539212A (en) * | 2012-01-29 | 2012-07-04 | 山东理工大学 | Manufacture process for section of skin placoid scale of cartilaginous fish through adopting metallographic sample method |
| CN102805664A (en) * | 2012-08-27 | 2012-12-05 | 王文艳 | Method for displaying and acquiring innervation image in human body levator ani muscle |
| CN103534571A (en) * | 2011-03-16 | 2014-01-22 | 卡利亚里大学 | Method, composition and kit of visualization and characterization of the nervous system by combining the staining for metal impregnation and immunohistochemistry |
| CN104089807A (en) * | 2014-07-07 | 2014-10-08 | 中国农业大学 | Germ staining method |
| CN104198256A (en) * | 2014-08-29 | 2014-12-10 | 温州医科大学附属第二医院 | Chemical staining method using niacinamide adenine dinucleotide diaphorase tissue |
| CN104749010A (en) * | 2015-03-31 | 2015-07-01 | 东华大学 | Preparation method of TEM (transmission electron microscope) ultrathin section samples of phosphorus-accumulating bacteria |
| US20150222877A1 (en) * | 2006-12-20 | 2015-08-06 | The Board Of Trustees Of The Leland Stanford Junior University | Arrangement and imaging of biological samples |
| CN106918484A (en) * | 2017-03-28 | 2017-07-04 | 武汉瑞福宁科技有限公司 | A kind of construction method of the threedimensional model based on histotomy |
| CN108508016A (en) * | 2018-04-09 | 2018-09-07 | 扬州大学附属医院 | A kind of prostate cancer three-dimensional pathology configuration method |
| CN108645683A (en) * | 2018-05-16 | 2018-10-12 | 绍兴文理学院 | A kind of palmitin fixer and its application |
| CN109085046A (en) * | 2018-10-25 | 2018-12-25 | 南京林业大学 | A kind of preparation method of leaf of bamboo internal anatomy 3 dimensional drawing |
| CN109859304A (en) * | 2018-11-16 | 2019-06-07 | 华中科技大学同济医学院附属同济医院 | Three-dimensional printing technology establishes the application in three-dimensional structure digital model in corneal limbal tissue in vitro |
| CN109887091A (en) * | 2019-02-27 | 2019-06-14 | 华侨大学 | A method of for lower limb and Thoracolumbar disk fascia to be redeveloped into 3D model |
| CN112393962A (en) * | 2020-11-30 | 2021-02-23 | 四川养麝研究所 | Three-dimensional reconstruction method of forest musk gland tissue |
| CN114113084A (en) * | 2021-11-11 | 2022-03-01 | 福州迈新生物技术开发有限公司 | Real-time accompanying film reading method based on digital pathological image and storage device |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106979878B (en) * | 2017-05-09 | 2023-12-15 | 贵阳德漫多医疗科技有限公司 | Histopathological specimen processing system |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2005007233A2 (en) * | 2003-06-20 | 2005-01-27 | Massachusetts Institute Of Technology | Application of electrical stimulation for functional tissue engineering in vitro and in vivo |
| CN1846614A (en) * | 2005-04-04 | 2006-10-18 | 沈渊瑶 | Method of constructing 3D tissue image |
| US20080013815A1 (en) * | 2006-06-16 | 2008-01-17 | Ruggero Scorcioni | Arborization Reconstruction |
| CN101246159A (en) * | 2008-03-07 | 2008-08-20 | 华中科技大学 | Method for fast identifying peripheral nerve bundle nature |
-
2010
- 2010-07-07 CN CN2010102191679A patent/CN101915693B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2005007233A2 (en) * | 2003-06-20 | 2005-01-27 | Massachusetts Institute Of Technology | Application of electrical stimulation for functional tissue engineering in vitro and in vivo |
| CN1846614A (en) * | 2005-04-04 | 2006-10-18 | 沈渊瑶 | Method of constructing 3D tissue image |
| US20080013815A1 (en) * | 2006-06-16 | 2008-01-17 | Ruggero Scorcioni | Arborization Reconstruction |
| CN101246159A (en) * | 2008-03-07 | 2008-08-20 | 华中科技大学 | Method for fast identifying peripheral nerve bundle nature |
Non-Patent Citations (4)
| Title |
|---|
| 《European Journal of Radiology》 20100531 Alexandra Borges et al. Imaging the trigeminal nerve 323-340 1-2 第74卷, 第2期 * |
| 《Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology, and Endodontology》 20050721 Tatsurou Tanaka et al. Utility of magnetic resonance cisternography using three-dimensional fast asymmetric spin-echo sequences with multiplanar reconstruction: The evaluation of sites of neurovascular compression of the trigeminal nerve 215-225 1-2 第100卷, 第2期 * |
| 《新疆医科大学学报》 20090831 吐尔逊江·达地汗等 人胚胎三叉神经运动纤维的定位研究 1043-1044，1048 1-2 第32卷, 第8期 * |
| 《现代生物医学进展》 20091231 吴励等 人胚胎三叉神经节细胞电镜及光镜观察 881-883 1-2 第9卷, 第5期 * |
Cited By (25)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US9578306B2 (en) * | 2006-12-20 | 2017-02-21 | The Board Of Trustees Of The Leland Stanford Junior University | Arrangement and imaging of biological samples |
| US20150222877A1 (en) * | 2006-12-20 | 2015-08-06 | The Board Of Trustees Of The Leland Stanford Junior University | Arrangement and imaging of biological samples |
| CN103534571B (en) * | 2011-03-16 | 2016-02-10 | 卡利亚里大学 | Manifest by the dyeing of combination metal impregnation and immunohistochemical staining and characterize neural method, composition and kit |
| CN103534571A (en) * | 2011-03-16 | 2014-01-22 | 卡利亚里大学 | Method, composition and kit of visualization and characterization of the nervous system by combining the staining for metal impregnation and immunohistochemistry |
| CN102243154A (en) * | 2011-04-02 | 2011-11-16 | 中山大学附属第一医院 | Method for dyeing peripheral nerves slice |
| CN102243154B (en) * | 2011-04-02 | 2016-05-11 | 中山大学附属第一医院 | A kind of method of dyeing peripheral nerves slice |
| CN102519774B (en) * | 2011-11-16 | 2014-02-12 | 中国水产科学研究院东海水产研究所 | Dyeing method for peripheral nerves of fish larvae and juveniles |
| CN102519774A (en) * | 2011-11-16 | 2012-06-27 | 中国水产科学研究院东海水产研究所 | Dyeing method for peripheral nerves of fish larvae and juveniles |
| CN102539212B (en) * | 2012-01-29 | 2013-08-21 | 山东理工大学 | Manufacture process for section of skin placoid scale of cartilaginous fish through adopting metallographic sample method |
| CN102539212A (en) * | 2012-01-29 | 2012-07-04 | 山东理工大学 | Manufacture process for section of skin placoid scale of cartilaginous fish through adopting metallographic sample method |
| CN102805664A (en) * | 2012-08-27 | 2012-12-05 | 王文艳 | Method for displaying and acquiring innervation image in human body levator ani muscle |
| CN104089807A (en) * | 2014-07-07 | 2014-10-08 | 中国农业大学 | Germ staining method |
| CN104198256A (en) * | 2014-08-29 | 2014-12-10 | 温州医科大学附属第二医院 | Chemical staining method using niacinamide adenine dinucleotide diaphorase tissue |
| CN104749010A (en) * | 2015-03-31 | 2015-07-01 | 东华大学 | Preparation method of TEM (transmission electron microscope) ultrathin section samples of phosphorus-accumulating bacteria |
| CN106918484A (en) * | 2017-03-28 | 2017-07-04 | 武汉瑞福宁科技有限公司 | A kind of construction method of the threedimensional model based on histotomy |
| CN108508016A (en) * | 2018-04-09 | 2018-09-07 | 扬州大学附属医院 | A kind of prostate cancer three-dimensional pathology configuration method |
| CN108508016B (en) * | 2018-04-09 | 2023-08-22 | 扬州大学附属医院 | Three-dimensional pathological configuration method for prostate cancer |
| CN108645683A (en) * | 2018-05-16 | 2018-10-12 | 绍兴文理学院 | A kind of palmitin fixer and its application |
| CN109085046A (en) * | 2018-10-25 | 2018-12-25 | 南京林业大学 | A kind of preparation method of leaf of bamboo internal anatomy 3 dimensional drawing |
| CN109859304A (en) * | 2018-11-16 | 2019-06-07 | 华中科技大学同济医学院附属同济医院 | Three-dimensional printing technology establishes the application in three-dimensional structure digital model in corneal limbal tissue in vitro |
| CN109859304B (en) * | 2018-11-16 | 2023-08-04 | 华中科技大学同济医学院附属同济医院 | Three-dimensional printing technology for establishing three-dimensional structure digital model outside cornea limbal tissue |
| CN109887091A (en) * | 2019-02-27 | 2019-06-14 | 华侨大学 | A method of for lower limb and Thoracolumbar disk fascia to be redeveloped into 3D model |
| CN112393962A (en) * | 2020-11-30 | 2021-02-23 | 四川养麝研究所 | Three-dimensional reconstruction method of forest musk gland tissue |
| CN114113084A (en) * | 2021-11-11 | 2022-03-01 | 福州迈新生物技术开发有限公司 | Real-time accompanying film reading method based on digital pathological image and storage device |
| WO2023082650A1 (en) * | 2021-11-11 | 2023-05-19 | 福州迈新生物技术开发有限公司 | Real-time concomitant slice reading method based on digital pathological image, and storage device |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101915693B (en) | 2012-07-25 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101915693B (en) | Human embryonic trigeminus based three-dimensional reconstruction method by using slide stainer | |
| CN107798979A (en) | A kind of method that 3D replicates printing skull, cranial nerve, brain tissue and brain vessel model | |
| DE102008015633B4 (en) | Perfusable bioreactor for the production and / or cultivation of a human or animal blood vessel and / or a human or animal tissue | |
| US20190385751A1 (en) | Disease condition information research method and system, and storage medium | |
| CN104568542A (en) | Method for fixing fish brain tissue specimen through systemic heart perfusion | |
| CN104287865B (en) | A kind of animal spinal cord living imaging Orientation observation device | |
| CN202078302U (en) | Portable tongue picture acquisition device | |
| CN2503554Y (en) | Artificial model of human body | |
| CN204542391U (en) | A kind of inspection of the tongue support | |
| CN201200396Y (en) | Image expression of hand section imaging of human body disease information and measuring equipment | |
| Li | Ultrasound biomicroscopy diagnosis analysis and fine care of anterior segment injury of traumatic anterior chamber based on intelligent virtual reality technology | |
| Coelho-Santos et al. | Reinforced thinned-skull window for repeated imaging of the neonatal mouse brain | |
| CN101986834A (en) | Method for preparing rhesus focal cerebral ischemia model | |
| RU78657U1 (en) | DEVICE FOR TRAINING NEURAL SURGEON DOCTORS WITH ENDOSCOPIC MANUAL SKILLS OF OPERATIVE INTERVENTIONS AT CLINICAL HEMATOMAS IN THE PROJECTION OF THE ANTERIAL CRANULAR FOSSAL | |
| Liu et al. | Nursing research of optic canal decompression operation under nasal endoscopic medical treatment based on intelligent internet of things for traumatic vision disorders | |
| Вовк et al. | Diaphonization as a method of modern morphological research | |
| CN206349059U (en) | The cranium brain model of liquid circulation dynamical system can be connected | |
| CN202920398U (en) | Multifunctional oral cavity treating device with camera | |
| RU125461U1 (en) | REFINING CAP | |
| CN203468569U (en) | Odontoscope with camera shooting function | |
| CN212847234U (en) | Cardiovascular intervention operation training simulator | |
| CN212261341U (en) | Neonate's umbilical cord changes display recorder | |
| Hu | Anatomy and physiology of bone perfusion in living and foss il birds as assessed by CT-scann ing, microsphere distribution, vascular contrast imaging and foramen measurement | |
| CN101755729B (en) | Human visual conduction path, blood supply integrated display sample and manufacturing method thereof | |
| TW201117111A (en) | Fish body image analysis system for use in the screening of a candidate as a skin whitening agent and applications of the same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20120725 Termination date: 20150707 |
|
| EXPY | Termination of patent right or utility model |