CN107084984A - A kind of image measuring method of myofibroblasts parameter - Google Patents
A kind of image measuring method of myofibroblasts parameter Download PDFInfo
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- CN107084984A CN107084984A CN201710252244.2A CN201710252244A CN107084984A CN 107084984 A CN107084984 A CN 107084984A CN 201710252244 A CN201710252244 A CN 201710252244A CN 107084984 A CN107084984 A CN 107084984A
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- 210000000651 myofibroblast Anatomy 0.000 title claims abstract description 32
- 238000000034 method Methods 0.000 title claims abstract description 17
- 210000003205 muscle Anatomy 0.000 claims abstract description 46
- 239000000835 fiber Substances 0.000 claims abstract description 45
- 238000005259 measurement Methods 0.000 claims abstract description 16
- 235000015277 pork Nutrition 0.000 claims abstract description 11
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 74
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical compound CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 claims description 27
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 23
- 235000019441 ethanol Nutrition 0.000 claims description 21
- 239000012188 paraffin wax Substances 0.000 claims description 21
- 239000001993 wax Substances 0.000 claims description 15
- 210000001519 tissue Anatomy 0.000 claims description 13
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 6
- 239000000975 dye Substances 0.000 claims description 6
- 238000002372 labelling Methods 0.000 claims description 6
- 238000000205 computational method Methods 0.000 claims description 4
- 229930040373 Paraformaldehyde Natural products 0.000 claims description 3
- 230000001476 alcoholic effect Effects 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 3
- 238000001035 drying Methods 0.000 claims description 3
- DZGCGKFAPXFTNM-UHFFFAOYSA-N ethanol;hydron;chloride Chemical compound Cl.CCO DZGCGKFAPXFTNM-UHFFFAOYSA-N 0.000 claims description 3
- 210000000630 fibrocyte Anatomy 0.000 claims description 3
- 235000011187 glycerol Nutrition 0.000 claims description 3
- HLUCICHZHWJHLL-UHFFFAOYSA-N hematein Chemical compound C12=CC=C(O)C(O)=C2OCC2(O)C1=C1C=C(O)C(=O)C=C1C2 HLUCICHZHWJHLL-UHFFFAOYSA-N 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 3
- 239000000155 melt Substances 0.000 claims description 3
- 230000007935 neutral effect Effects 0.000 claims description 3
- 229920002866 paraformaldehyde Polymers 0.000 claims description 3
- 238000010186 staining Methods 0.000 claims description 3
- 230000007704 transition Effects 0.000 claims description 3
- 230000000007 visual effect Effects 0.000 claims description 3
- 239000002023 wood Substances 0.000 claims description 3
- 239000008096 xylene Substances 0.000 claims description 3
- 239000003106 tissue adhesive Substances 0.000 claims 1
- 210000004027 cell Anatomy 0.000 abstract description 3
- 235000013372 meat Nutrition 0.000 description 3
- 239000000853 adhesive Substances 0.000 description 2
- 230000001070 adhesive effect Effects 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 241000208340 Araliaceae Species 0.000 description 1
- 235000005035 Panax pseudoginseng ssp. pseudoginseng Nutrition 0.000 description 1
- 235000003140 Panax quinquefolius Nutrition 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 235000008434 ginseng Nutrition 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 210000001087 myotubule Anatomy 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 238000009394 selective breeding Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/84—Systems specially adapted for particular applications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/36—Embedding or analogous mounting of samples
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- G—PHYSICS
- G06—COMPUTING; CALCULATING OR COUNTING
- G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
- G06T7/00—Image analysis
- G06T7/0002—Inspection of images, e.g. flaw detection
- G06T7/0004—Industrial image inspection
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- G—PHYSICS
- G06—COMPUTING; CALCULATING OR COUNTING
- G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
- G06T2207/00—Indexing scheme for image analysis or image enhancement
- G06T2207/10—Image acquisition modality
- G06T2207/10056—Microscopic image
- G06T2207/10061—Microscopic image from scanning electron microscope
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Abstract
The invention discloses a kind of image measuring method of myofibroblasts parameter, in the present invention, we are after the muscle fibre sample of pork is prepared for, micro-image is gathered, then by image processing algorithm, program of writing carrys out automatic Ground Split and goes out myofibroblasts, and the calculating of the parameters such as area, the diameter of individual cells is completed on this basis, so as to realize the automatic measurement of pork muscle fibre parameter, the workload of measurement is reduced, the degree of accuracy of measurement is improved.
Description
Technical field
The invention belongs to detection field, more particularly to a kind of image measuring method of myofibroblasts parameter.
Background technology
It can be used for evaluating different raising or growth environment to the quantitative analysis of meat fiber for the influence of muscle, also may be used
For work such as seed selections.Existing analysis method still relies on manual measurement substantially, i.e., by people come under viewing microscope
Muscle sample, and being measured accordingly, has that the amount of labour is big, inefficiency, accuracy.There is part document
In image processing software has been used to help to measure, but still dependent on artificial selection, do not have in terms of the amount of labour and operating efficiency
There is big change.
For example in paper《Different images analysis software quantitatively calculates muscle fiber types quantity and area》In, author uses
The image processing softwares such as ImageJ, Image Pro plus and Photoshop, by way of man-machine interactively, select flesh by hand
Fibrocyte, then calculates its area by image processing software.Although improving measurement using image processing software reference area
Accuracy, it is to avoid the past is estimated the error caused during the area of the myofibroblasts of irregular shape by people, but measurement
Still relied on process nature artificial.
In the present invention, we gather micro-image, then pass through image after the muscle fibre sample of pork is prepared for
Processing Algorithm, program of writing carrys out automatic Ground Split and goes out myofibroblasts, and completes area, the diameter of individual cells on this basis
Etc. the calculating of parameter, so as to realize the automatic measurement of pork muscle fibre parameter, the workload of measurement is reduced, the accurate of measurement is improved
Degree.
The content of the invention
It is an object of the invention to provide a kind of image measuring method of myofibroblasts parameter, the present invention by image at
The method of reason, can be automatically completed the measurement of pork muscle fibre parameter, and the amount of labour is small, and operating efficiency is high, as a result stablizes, accurately
Degree is high, and can avoid in artificial operation because of fatigue or the introduced error result of supervisor's factor of judgment.
To achieve the above object, technical scheme is as follows：
A kind of image measuring method of myofibroblasts parameter, comprises the following steps:
(1) image of muscle fibre and the scale of image are obtained；
(2) image measurement of muscle fibre parameter：
(2.1) the coloured image I for obtaining shooting is converted into gray level image G, and then entering row threshold division to G obtains two-value
Image B1；B1In, dark myofibroblasts region is black, and background area is white；
(2.2) to B1Connected component labeling is carried out, area is then removed and is less than given threshold value TA1White portion；Ask for white
The inverse image of bianry image after the removal of region, obtains B2；
(2.3) to B2Connected component labeling is carried out, mark image L is obtained1, then remove L1In welt region, marked
Image L2；
(2.4) L is removed2Middle area is less than given threshold value TA2Or more than given threshold value TA3Region, obtain mark image L3；
(2.5) to L3In each region, calculate its area and convex closure area ratio r, remove L3Middle r values are less than given threshold
Value TacarRegion, obtain mark image L4；
(2.6) to L4In each region, carry out holes filling, produce muscle fibre region；
(2.7) calculated according to muscle fibre region and obtain myofibroblasts parameter.
Further to improve, in step (2.7), the myofibroblasts parameter includes the real area of muscle fibre, and flesh is fine
The real area computational methods of dimension are as follows：
The relative area for calculating muscle fibre region is the pixel number A in muscle fibre regionI；According to given in image
Scale, calculates the real area A=A of the physical length s, then muscle fibre in image corresponding to 1 length in pixelsI·s2。
Further to improve, in step (2.7), the myofibroblasts parameter includes the actual diameter of muscle fibre, and flesh is fine
The actual diameter computational methods of dimension are as follows：
The profile in muscle fibre region is extracted, the distance between each pair dissimilarity on profile is calculated, with all dissimilaritys pair
The maximum of distance is the diameter d of muscle fibreI, unit is pixel；According to the scale given in image, 1 pixel in image is calculated long
Degree corresponding physical length s, then muscle fibre actual diameter d=dI·s。
It is further to improve, at least one in welt region even some myofibroblasts region in step (2.3)
Pixel falls the region outside the most upper of the outer rim in image, i.e. image, most descending, left column or right column, then this muscle fibre
Cell is in image L1Interior region is referred to as welt region.
Further to improve, the myofibroblasts are pork fibrocyte.
Further to improve, in step (1), the image of muscle fibre shoots the sample micro-image in multiple visuals field by camera
Obtain.
Further to improve, the sample is made by following steps：
(1.1) it is fixed：With 4% paraformaldehyde fixed sample 24 hours；
(1.2) it is dehydrated：Sample after will be washed is put into the alcohol for 70%, 80%, 90%, 100%, 100% respectively
By staying overnight, soak to slough the moisture in sample within 8 hours, 8 hours, 1 hour, 1 hour；
(1.3) it is transparent：With 1:1 dimethylbenzene:Absolute alcohol soaks 1 hour, then with xylene soak 1 hour, to replace
The absolute alcohol gone out in tissue block；
(1.4) waxdip and embedding：Transparent good sample is put into 1:1 dimethylbenzene:Transition 30 minutes in paraffin, then by its
It is put into the wax cup equipped with paraffin 30 minutes, will need to be carried out in incubator after melted paraffin wax, Temperature of Warm Case must not be higher than 56 DEG C, will
The tissue block for having soaked wax is put into the carton for filling melted paraffin, tissue block is embedded in paraffin, is placed in basin rapidly, is treated
It is standby after cooling down；
(1.5) cut into slices：The thin of 6 μ m-thicks is cut into slicer after the paraffin embedded tissues made are adhered into platform wood after repairing and cutting
Piece, thin slice is continuously into wax banding；
(1.6) exhibition piece and bonding die：The wax band being cut into is opened up in 30 DEG C of warm water, then with 1:1 egg:Glycerine is mixed
Agent is adhesive, and tissue slice is adhered on slide, then is placed on drying in 37 DEG C of incubators, no less than 24h；
(1.7) dewax rehydration：Water-bath pot temperature is adjusted to 60 DEG C, when water temperature control is at 60 DEG C, will be cut into slices together with section
Frame is put into a dry staining jar, is put into water-bath, is closed the lid, and 30min to wax melts；
(1.8) paraffin section is then placed in 100%, 95%, 90%, 80%, 70% through each 5min of dewaxing of dimethylbenzene I, II
Each 3-5min in the alcoholic solution of graded concentrations, places into 3min in distilled water；
(1.9) dye：Section, which is put into haematine, dyes 15min；
(1.10) wash：15min is rinsed with running water flowing water；Section color is set to become blue；
(1.11) break up：Section is put into 1% hydrochloride ethanol liquid and faded, 2 seconds to several tens of seconds；See that section reddens, face
Color is shallower；
(1.12) rinse：Section, which is placed into, makes it recover blueness in running water flowing water；
(1.13) it is dehydrated I：Each 3-5min in the ethanol of 50% ethanol → 70% of cutting into slices ethanol → 80%；
(1.14) redye：With 0.5% Yihong ethanol counterstain 1min；
(1.15) it is dehydrated II：Section is put into 95% ethanol and washes away unnecessary red, 3- in absolute ethyl alcohol is then placed in
5min；Unnecessary ethanol is finally blotted with blotting paper；
(1.16) it is transparent：Section is put into each 3-5min in dimethylbenzene I, II；
(1.17) envelope is hidden：Neutral gum is sealed up for safekeeping.
Embodiment
Embodiment
Institute's extracting method of the present invention includes two Main Stages：The preparation of pork muscle sample and image-acquisition phase, and
Image procossing and measuring phases.
(1) sample preparation is obtained with image
(1.1) it is fixed：With 4% paraformaldehyde fixed sample 24 hours.
(1.2) it is dehydrated：Sample after will be washed is put into as 70%, 80%, 90%, 100% (absolute alcohol), 100% (nothing
Water-alcohol) alcohol in respectively through overnight, soak to slough the moisture in sample within 8 hours, 8 hours, 1 hour, 1 hour.
(1.3) it is transparent：With 1:1 dimethylbenzene:Absolute alcohol soaks 1 hour, then with xylene soak 1 hour, to replace
The absolute alcohol gone out in tissue block.
(1.4) waxdip and embedding：Transparent good sample is put into 1:1 dimethylbenzene:Transition 30 minutes in paraffin, then by its
It is put into the wax cup equipped with paraffin 30 minutes (will need to be carried out in incubator after melted paraffin wax, Temperature of Warm Case must not be higher than 56 DEG C).
The tissue block for having soaked wax is put into the carton for filling melted paraffin, tissue block is embedded in paraffin, is placed in basin rapidly,
After standby after its cooling.
(1.5) cut into slices：The thin of 6 μ m-thicks is cut into slicer after the paraffin embedded tissues made are adhered into platform wood after repairing and cutting
Piece, thin slice should be continuously into wax banding.
(1.6) exhibition piece and bonding die：The wax band being cut into is opened up in 30 DEG C of warm water, then with 1:1 egg:Glycerine is mixed
Agent is adhesive, and tissue slice is adhered on slide, then is placed on drying in 37 DEG C of incubators (being no less than 24h).
(1.7) dewax rehydration：Water-bath pot temperature is adjusted to 60 DEG C, when water temperature control is at 60 DEG C, will be cut into slices together with section
Frame is put into a dry staining jar, is put into water-bath, and close the lid (salable), and 30min to wax melts.
(1.8) paraffin section is then placed in 100%, 95%, 90%, 80%, 70% through each 5min of dewaxing of dimethylbenzene I, II
Each 3-5min in the alcoholic solution of graded concentrations, places into 3min in distilled water.
(1.9) dye：Section, which is put into haematine, dyes about 15min.
(1.10) wash：About 15min is rinsed with running water flowing water.Make section color become blue (or to be put into alkaline water
Can), but it is noted that flowing water can not be excessive, come off to prevent section.
(1.11) break up：Section is put into 1% hydrochloride ethanol liquid and faded, about 2 seconds to several tens of seconds.See that section reddens,
Color is shallower.
(1.12) rinse：Section, which is placed into, makes it recover blueness in running water flowing water.
(1.13) it is dehydrated I：Each 3-5min in the ethanol of 50% ethanol → 70% of cutting into slices ethanol → 80%.
(1.14) redye：With 0.5% Yihong ethanol counterstain 1min.
(1.15) it is dehydrated II：Section is put into 95% ethanol and washes away unnecessary red, 3- in absolute ethyl alcohol is then placed in
5min.Unnecessary ethanol is finally blotted with blotting paper.
(1.16) it is transparent：Section is put into each 3-5min in dimethylbenzene I, II.
(1.17) envelope is hidden：Neutral gum is sealed up for safekeeping
(1.18) image is obtained：Sample is placed under microscope, the sample micro-image in multiple visuals field is shot by camera,
Scale should be added in image.
(2) image measurement of muscle fibre parameter
(2.1) the coloured image I for obtaining shooting is converted into gray level image G, and then entering row threshold division to G obtains two-value
Image B1。B1In, dark myofibroblasts region is black, and background area is white；
(2.2) to B1Connected component labeling is carried out, area is then removed and is less than given threshold value TA1Those white portions.Ask for
The inverse image of bianry image after the removal of region, obtains B2；
(2.3) to B2Connected component labeling is carried out, mark image L is obtained1, then remove L1In welt region, i.e., in region
At least one pixel falls in the outer rim (i.e. the highest and lowest row and most left and right column of image) of image, obtains
Mark image L2；Welt region is to refer to indivedual myofibroblasts to be not all of in image L1In the range of, but a part is in image
L1Interior, another part is in image L1Outside, in image L1The incomplete myofibroblasts region of interior part is described welt area
Domain.
(2.4) L is removed2Middle area is less than given threshold value TA2Or more than given threshold value TA3Region, obtain mark image L3；
(2.5) to L3In each region, calculate its area and convex closure area ratio r, remove L3Middle r values are less than given threshold
Value TacarRegion, obtain mark image L4；
(2.6) to L4In each region, holes filling is carried out to it, the muscle fibre region of pork is produced；
(2.7) area (the i.e. pixel number) A in muscle fibre region is calculatedI；
(2.8) profile in muscle fibre region is extracted, the distance between each pair dissimilarity on profile is calculated, with all these points
To distance maximum be muscle fibre diameter (in units of pixel) dI；
(2.9) scale according to given in image, calculates the physical length s corresponding to 1 length in pixels in image, then flesh
The real area of fiber is A=AI·s2, actual diameter is d=dI·s。
Carried although the present invention is the muscle fibre parameter measurement for being directed to pork, if the muscle fibre ginseng of other meats
Number measurement, if the muscle fibre form of these meats is similar with pork, the method for the present invention is equally applicable, therefore should be regarded as
The scope that the present invention is covered.
Signified muscle fibre parameter of the invention, refers mainly to the area and diameter measurement in muscle fibre cross section.
As described above, the embodiment of the present invention is described in detail, but as long as essentially without this hair of disengaging
Bright the characteristics of and effect, can there is many variations, and this will be readily apparent to persons skilled in the art, therefore,
The example of these deformations is also integrally incorporated within protection scope of the present invention.
Claims (7)
1. a kind of image measuring method of myofibroblasts parameter, it is characterised in that comprise the following steps:
(1) image of muscle fibre and the scale of image are obtained；
(2) image measurement of muscle fibre parameter：
(2.1) the coloured image I for obtaining shooting is converted into gray level image G, and then entering row threshold division to G obtains bianry image
B1；B1In, dark myofibroblasts region is black, and background area is white；
(2.2) to B1Connected component labeling is carried out, area is then removed and is less than given threshold value TA1White portion；Ask for white portion
The inverse image of bianry image after removal, obtains B2；
(2.3) to B2Connected component labeling is carried out, mark image L is obtained1, then remove L1In welt region, obtain mark image
L2；
(2.4) L is removed2Middle area is less than given threshold value TA2Or more than given threshold value TA3Region, obtain mark image L3；
(2.5) to L3In each region, calculate its area and convex closure area ratio r, remove L3Middle r values are less than given threshold value
TacarRegion, obtain mark image L4；
(2.6) to L4In each region, carry out holes filling, produce muscle fibre region；
(2.7) calculated according to muscle fibre region and obtain myofibroblasts parameter.
2. the image measuring method of myofibroblasts parameter as claimed in claim 1, it is characterised in that in step (2.7), institute
Stating myofibroblasts parameter includes the real area of muscle fibre, and the real area computational methods of muscle fibre are as follows：
The relative area for calculating muscle fibre region is the pixel number A in muscle fibre regionI；Scale according to given in image,
Calculate the real area A=A of the physical length s, then muscle fibre in image corresponding to 1 length in pixelsI·s2。
3. the image measuring method of myofibroblasts parameter as claimed in claim 1, it is characterised in that in step (2.7), institute
Stating myofibroblasts parameter includes the actual diameter of muscle fibre, and the actual diameter computational methods of muscle fibre are as follows：
The profile in muscle fibre region is extracted, the distance between each pair dissimilarity on profile is calculated, with the distance of all dissimilaritys pair
Maximum be muscle fibre diameter dI, unit is pixel；According to the scale given in image, 1 length in pixels institute in image is calculated
Corresponding physical length s, then muscle fibre actual diameter d=dI·s。
4. the image measuring method of myofibroblasts parameter as claimed in claim 1, it is characterised in that in step (2.3)
At least one pixel falls the outer rim in image, i.e. image most in even some the myofibroblasts region of welt region
Region outside upper, most descending, left column or right column, then this myofibroblasts is in image L1Interior region is referred to as welt region.
5. the image measuring method of myofibroblasts parameter as claimed in claim 1, it is characterised in that the myofibroblasts
For pork fibrocyte.
6. the image measuring method of myofibroblasts parameter as claimed in claim 1, it is characterised in that in step (1), flesh is fine
The sample micro-image that the image of dimension shoots multiple visuals field by camera is obtained.
7. the image measuring method of myofibroblasts parameter as claimed in claim 6, it is characterised in that the sample by with
Lower step is made：
(1.1) it is fixed：With 4% paraformaldehyde fixed sample 24 hours；
(1.2) it is dehydrated：Will it is washed after sample be put into the alcohol for 70%, 80%, 90%, 100%, 100% respectively through
Overnight, soak to slough the moisture in sample within 8 hours, 8 hours, 1 hour, 1 hour；
(1.3) it is transparent：With 1:1 dimethylbenzene:Absolute alcohol soaks 1 hour, then with xylene soak 1 hour, to displace group
Knit the absolute alcohol in block；
(1.4) waxdip and embedding：Transparent good sample is put into 1:1 dimethylbenzene:Transition 30 minutes in paraffin, then put it into
30 minutes in wax cup equipped with paraffin, it will need to be carried out in incubator after melted paraffin wax, Temperature of Warm Case must not be higher than 56 DEG C, will soak
The tissue block of wax is put into the carton for filling melted paraffin, tissue block is embedded in paraffin, is placed in basin rapidly, treats that its is cold
But it is standby after；
(1.5) cut into slices：The paraffin embedded tissues made are adhered to the thin slice for being cut into 6 μ m-thicks after platform wood with slicer after repairing and cutting, it is thin
Piece is continuous into wax banding；
(1.6) exhibition piece and bonding die：The wax band being cut into is opened up in 30 DEG C of warm water, then with 1:1 egg:Glycerine intermixture is
Adhesive, tissue slice is adhered on slide, then is placed on drying in 37 DEG C of incubators, no less than 24h；
(1.7) dewax rehydration：Water-bath pot temperature is adjusted to 60 DEG C, when water temperature control is at 60 DEG C, section put together with slide holding frame
Enter in a dry staining jar, be put into water-bath, close the lid, 30min to wax melts；
(1.8) paraffin section is then placed in 100%, 95%, 90%, 80%, 70% at different levels through each 5min of dewaxing of dimethylbenzene I, II
Each 3-5min in the alcoholic solution of concentration, places into 3min in distilled water；
(1.9) dye：Section, which is put into haematine, dyes 15min；
(1.10) wash：15min is rinsed with running water flowing water；Section color is set to become blue；
(1.11) break up：Section is put into 1% hydrochloride ethanol liquid and faded, 2 seconds to several tens of seconds；See that section reddens, color compared with
It is shallow；
(1.12) rinse：Section, which is placed into, makes it recover blueness in running water flowing water；
(1.13) it is dehydrated I：Each 3-5min in the ethanol of 50% ethanol → 70% of cutting into slices ethanol → 80%；
(1.14) redye：With 0.5% Yihong ethanol counterstain 1min；
(1.15) it is dehydrated II：Section is put into 95% ethanol and washes away unnecessary red, 3-5min in absolute ethyl alcohol is then placed in；
Unnecessary ethanol is finally blotted with blotting paper；
(1.16) it is transparent：Section is put into each 3-5min in dimethylbenzene I, II；
(1.17) envelope is hidden：Neutral gum is sealed up for safekeeping.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710252244.2A CN107084984A (en) | 2017-04-18 | 2017-04-18 | A kind of image measuring method of myofibroblasts parameter |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710252244.2A CN107084984A (en) | 2017-04-18 | 2017-04-18 | A kind of image measuring method of myofibroblasts parameter |
Publications (1)
| Publication Number | Publication Date |
|---|---|
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| CN109975294A (en) * | 2019-04-03 | 2019-07-05 | 天津市畜牧兽医研究所 | A kind of measuring method of musculature fiber sum |
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