CN108956243A - A kind of palmitin fixer and preparation method thereof - Google Patents
A kind of palmitin fixer and preparation method thereof Download PDFInfo
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- CN108956243A CN108956243A CN201810468473.2A CN201810468473A CN108956243A CN 108956243 A CN108956243 A CN 108956243A CN 201810468473 A CN201810468473 A CN 201810468473A CN 108956243 A CN108956243 A CN 108956243A
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- PVNIQBQSYATKKL-UHFFFAOYSA-N Glycerol trihexadecanoate Natural products CCCCCCCCCCCCCCCC(=O)OCC(OC(=O)CCCCCCCCCCCCCCC)COC(=O)CCCCCCCCCCCCCCC PVNIQBQSYATKKL-UHFFFAOYSA-N 0.000 title claims abstract description 44
- 238000002360 preparation method Methods 0.000 title claims abstract description 16
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 78
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 claims abstract description 72
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 claims abstract description 31
- 239000000787 lecithin Substances 0.000 claims abstract description 31
- 229940067606 lecithin Drugs 0.000 claims abstract description 31
- 235000010445 lecithin Nutrition 0.000 claims abstract description 31
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 29
- 229960000935 dehydrated alcohol Drugs 0.000 claims abstract description 20
- 239000002994 raw material Substances 0.000 claims abstract description 18
- 239000000463 material Substances 0.000 claims abstract description 14
- 238000003756 stirring Methods 0.000 claims abstract description 12
- 230000015572 biosynthetic process Effects 0.000 claims abstract description 11
- WSFSSNUMVMOOMR-NJFSPNSNSA-N methanone Chemical compound O=[14CH2] WSFSSNUMVMOOMR-NJFSPNSNSA-N 0.000 claims abstract description 9
- 238000002156 mixing Methods 0.000 claims abstract description 8
- 229940022682 acetone Drugs 0.000 claims abstract description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 96
- RBTBFTRPCNLSDE-UHFFFAOYSA-N 3,7-bis(dimethylamino)phenothiazin-5-ium Chemical compound C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 RBTBFTRPCNLSDE-UHFFFAOYSA-N 0.000 claims description 24
- 229960000907 methylthioninium chloride Drugs 0.000 claims description 24
- UYVVLXVBEQAATF-UHFFFAOYSA-N 4-(1,3,7,12-tetrahydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1h-cyclopenta[a]phenanthren-17-yl)pentanoic acid Chemical compound OC1CC2CC(O)CC(O)C2(C)C2C1C1CCC(C(CCC(O)=O)C)C1(C)C(O)C2 UYVVLXVBEQAATF-UHFFFAOYSA-N 0.000 claims description 4
- 239000000843 powder Substances 0.000 claims description 4
- 239000007788 liquid Substances 0.000 claims description 3
- 241000233855 Orchidaceae Species 0.000 claims 2
- 230000003796 beauty Effects 0.000 claims 2
- 210000001165 lymph node Anatomy 0.000 abstract description 46
- 230000000694 effects Effects 0.000 abstract description 21
- 206010028980 Neoplasm Diseases 0.000 abstract description 14
- 201000011510 cancer Diseases 0.000 abstract description 13
- 238000002224 dissection Methods 0.000 abstract description 8
- 238000003364 immunohistochemistry Methods 0.000 abstract description 7
- 210000004027 cell Anatomy 0.000 abstract description 5
- 208000007433 Lymphatic Metastasis Diseases 0.000 abstract description 3
- 238000012360 testing method Methods 0.000 abstract description 3
- 229960004279 formaldehyde Drugs 0.000 abstract description 2
- 239000002953 phosphate buffered saline Substances 0.000 description 36
- 239000000243 solution Substances 0.000 description 34
- 210000001519 tissue Anatomy 0.000 description 16
- 238000012545 processing Methods 0.000 description 14
- 238000004043 dyeing Methods 0.000 description 9
- 210000000577 adipose tissue Anatomy 0.000 description 8
- 230000000295 complement effect Effects 0.000 description 8
- 239000000470 constituent Substances 0.000 description 8
- 238000000034 method Methods 0.000 description 6
- 239000000203 mixture Substances 0.000 description 6
- 238000011282 treatment Methods 0.000 description 6
- 238000004090 dissolution Methods 0.000 description 5
- 239000000975 dye Substances 0.000 description 5
- 235000019441 ethanol Nutrition 0.000 description 5
- 230000007935 neutral effect Effects 0.000 description 5
- 206010021703 Indifference Diseases 0.000 description 4
- 230000008859 change Effects 0.000 description 4
- 238000005238 degreasing Methods 0.000 description 4
- 239000004615 ingredient Substances 0.000 description 4
- 210000004072 lung Anatomy 0.000 description 4
- 238000002844 melting Methods 0.000 description 4
- 230000008018 melting Effects 0.000 description 4
- 208000011645 metastatic carcinoma Diseases 0.000 description 4
- 206010061289 metastatic neoplasm Diseases 0.000 description 4
- 230000007170 pathology Effects 0.000 description 4
- 238000011470 radical surgery Methods 0.000 description 4
- 206010006187 Breast cancer Diseases 0.000 description 3
- 208000026310 Breast neoplasm Diseases 0.000 description 3
- 210000004953 colonic tissue Anatomy 0.000 description 3
- 229960004756 ethanol Drugs 0.000 description 3
- 238000011532 immunohistochemical staining Methods 0.000 description 3
- 210000005075 mammary gland Anatomy 0.000 description 3
- 238000005070 sampling Methods 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 238000012546 transfer Methods 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- 101710114425 Homeobox protein Nkx-2.1 Proteins 0.000 description 2
- 102100027893 Homeobox protein Nkx-2.1 Human genes 0.000 description 2
- 206010027476 Metastases Diseases 0.000 description 2
- 101710088547 Thyroid transcription factor 1 Proteins 0.000 description 2
- 101710159262 Transcription termination factor 1 Proteins 0.000 description 2
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- 239000012530 fluid Substances 0.000 description 2
- 230000000762 glandular Effects 0.000 description 2
- 238000007689 inspection Methods 0.000 description 2
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 2
- 210000002751 lymph Anatomy 0.000 description 2
- 230000001926 lymphatic effect Effects 0.000 description 2
- 230000009401 metastasis Effects 0.000 description 2
- 230000000849 parathyroid Effects 0.000 description 2
- 238000004393 prognosis Methods 0.000 description 2
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- 102000007469 Actins Human genes 0.000 description 1
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- 108050005493 CD3 protein, epsilon/gamma/delta subunit Proteins 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 208000012661 Dyskinesia Diseases 0.000 description 1
- 101000897405 Homo sapiens B-lymphocyte antigen CD20 Proteins 0.000 description 1
- 101000994460 Homo sapiens Keratin, type I cytoskeletal 20 Proteins 0.000 description 1
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- 238000006243 chemical reaction Methods 0.000 description 1
- 210000000038 chest Anatomy 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000012321 colectomy Methods 0.000 description 1
- 210000001072 colon Anatomy 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000012937 correction Methods 0.000 description 1
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- 230000007812 deficiency Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000004907 gland Anatomy 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
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- 150000002576 ketones Chemical class 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 210000001365 lymphatic vessel Anatomy 0.000 description 1
- 230000013011 mating Effects 0.000 description 1
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 1
- 210000004877 mucosa Anatomy 0.000 description 1
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Classifications
-
- 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
-
- 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
- G01N2001/305—Fixative compositions
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
This application involves a kind of palmitin fixers and preparation method thereof, belong to test sample preparation technical field.Using formaldehyde, lecithin, dehydrated alcohol and acetone as raw material, after mixing evenly by dehydrated alcohol, formaldehyde and acetone, lecithin is added, it stirs and makes it dissolve, the concentration of above-mentioned each raw material is respectively as follows: 50 ~ 150ml/L of formaldehyde in the fixer of formation, 30 ~ 70g/L of lecithin, dehydrated alcohol 200 ~ 300 ml/L, 200 ~ 300ml/L of acetone.The application is applied to the preparation of palmitin fixer, fixation especially in cancer class sample in lymph node dissection materials, it not only ensure that the timely abundant fixation of lymph node, the dissection for reducing Pathologis is drawn materials the time, lymph node cells form is also ensured to save completely, without deformation, immunohistochemistry effect is good, effectively increases lymph nodes number and accurate lymphatic metastasis positive rate.
Description
Technical field
This application involves a kind of palmitin fixers and preparation method thereof, belong to test sample preparation technical field.
Background technique
The main path of cancer metastasis is lymphatic channel, and cancer cell is transferred to lymph node through lymphatic return, not only causes part
Disorganization, and hidden in lymph node, become the cradle of other Organ relative weights, excision includes primary tumor and drainage region
The radical surgery of lymph node is the mostly important operative treatment mode of cancer patient.Lymph node solution after cancer radical surgery
Cut open be with pathologic sampling clinic diagnosis one of important step.There is the sufficient lymph node radical excision of studies have shown that can be substantially
Degree reduces cancer-related events, even if all lymph node pathologies check without transfer, also has weight for the long-term survival of patient
It influences.The transfering state of lymph node is often the independent prognostic factor of cancer patient, and the lymph node number of excision is more, patient's
Recurrence rate and case fatality rate are lower, and this trend is also embodied in the patient of lymphatic metastasis.There is research to confirm, the lymph of transfer
Knot minimum diameter is 0.4cm, and whether there is or not shift to be difficult to judge only according to naked eyes general characteristics for lymph node.Obviously, it sufficiently and definitely drenches
The incidence for checking and can reducing lymph node false negative is fawned on, Axelsson etc. has checked the operation of 31679 patient with breast cancers
Sample prompts the lymph node of excision more, and the rate of transform of lymph node is higher；With with 10-14 pieces of lymph node materials be radix with
20 pieces of lymph node materials are compared as radix, and the Lymph node positive rate of diameter 1-5mm can be made to be increased to 25.9% by 14.2%, made
Diameter 11-20mm Lymph node positive rate is increased to 90.0% by 38.6%.Cancerous area lymph node resection number and transfer are positive
An important factor for rate is influence patient's prognosis, and one of the important evidence of therapeutic scheme is formulated, total lymph node materials are to true
Determine cancer staging, assessment prognosis and formulate individualized treatment scheme to be of great significance.
Currently, the lymph node dissection materials of cancer radical correction sample mostly continue to use conventional method, i.e., to fresh or formal
The fixed specimens from pri of woods, Pathology Doctors ' cut the method that hand is touched using knife, and rule of thumb judgement is usually wrapped in adipose tissue
Lymph node and materials.And most difficult during lymph node materials is exactly that adipose tissue is excessive, leads to acquirement lymph node numbers
Deficiency seriously affects the postoperative therapeutic scheme of patient so that clinicopathologic stage is inaccurate, and especially less than 0.5cm's is small
Lymph node easily missing inspection, and these small lymph nodes equally exist the possibility of metastatic carcinoma.Lymph node dissection materials quantity, quality with
And required time is influenced by many factors such as fat content, the anatomies even sense of responsibility of operator.Clinical practice work
It is badly in need of that the adipose tissue for interfering lymph node exposure and judgement can be dissolved in work, and can guarantee the sufficiently fixed method of lymph node.
This research is to solve the problems, such as this urgent clinical needs and design.
Although existing cancer pathology scholar points out that lymph node harvest may need to consider lipolyse technology, Recent study
Fat solvent is focused primarily upon in mammary cancer armpit lymph gland Radical dissection, to protect lymphatic vessel, prevents traditional armpit from cleaning hand
Paralysis brachialis, pain caused by art propose the complication such as shoulder dyskinesia, edema of the upper extremity, armpit and chest hypohydrops.About packet
Include specimen sampling after the clinical common cancer radical surgery including regular breast cancer modified skinsuture, stomach and Radical Colectomy for Carcinoma of Colon
Degreasing fixing means is rarely reported.Fat solvent in operation is applied to mammary gland routine modified skinsuture by Zhang Qinqin etc.
Fresh specimens, and with routinely dissect sampling method compared with, prompt Fat solvent pretreatment sample can increase lymph node inspection
Number, attenuating dissection difficulty and shortening dissection time out.But main problem existing for this method is not to be suitable for clinic
More common fixed preparation.
Based on this, the application is made.
Summary of the invention
For drawbacks described above present in existing lymph nodes technology, it is solid that the application provides a kind of completely new palmitin first
Determine liquid, which has not only had both the multiple action of fat melting, fixation, lymph node dyeing, in dissolution through processed rouge
While fat tissue, guarantee the complete and optimum dyeing effect of lymph node structure.
To achieve the above object, the technical solution that the application takes is as follows:
A kind of palmitin fixer, each component including pressing densimeter below: 50~150ml/L of formaldehyde, lecithin 30~
70g/L, 200~300ml/L of dehydrated alcohol, 200~300ml/L of acetone.
Further, as preferred:
It further include having deoxycholic aicd, methanol, PBS solution, concentration of the deoxycholic aicd in fixer is 30~70g/L, first
Concentration of the alcohol in fixer is 200-300ml/L, and it is 100-200ml/L that PBS solution, which is added to the concentration in fixer,.It is more excellent
Choosing, in the palmitin fixer composition, each component concentration is respectively as follows: formaldehyde 100ml/L, lecithin 50g/L, deoxycholic aicd
47.5g/L, dehydrated alcohol 250ml/L, methanol 250ml/L, acetone 250ml/L, PBS solution 1 ×.
Further include having deoxycholic aicd, methanol, PBS solution and methylene blue, concentration of the deoxycholic aicd in fixer be 30~
70g/L, concentration of the methanol in fixer are 200-300ml/L, and it is 100- that PBS solution, which is added to the concentration in fixer,
200ml/L, concentration of the methylene blue in fixer are 0.05-0.15g/L.It is furthermore preferred that in the palmitin fixer composition, each group
Point concentration is respectively as follows: formaldehyde 100ml/L, lecithin 50g/L, deoxycholic aicd 47.5g/L, dehydrated alcohol 250ml/L, methanol
250ml/L, acetone 250ml/L, PBS solution 1 ×, methylene blue 0.1g/L.
Above-mentioned PBS solution is that pbs powder is added to stirring and dissolving in distilled water to be formulated, or with 7 × PBS solution
Etc. high powers PBS solution dilute.
Meanwhile present invention also provides the preparation methods of palmitin fixer as characterized above, with formaldehyde, lecithin, nothing
Water-ethanol and acetone are as raw material, after mixing evenly by dehydrated alcohol, formaldehyde and acetone, lecithin are added, stirs and keeps its molten
Solution, the concentration of above-mentioned each raw material is respectively as follows: 50~150ml/L of formaldehyde in the fixer of formation, lecithin 30~70g/L, anhydrous
200~300ml/L of ethyl alcohol, 200~300ml/L of acetone.The raw material further includes having deoxycholic aicd, is added and goes after lecithin addition
Oxycholic acid is simultaneously stirred to dissolving, and deoxycholic aicd concentration is 30~70g/L in the fixer of formation.
Further, as preferred:
The raw material further includes having methanol, and methanol concentration is 200-300ml/L in the fixer of formation.The raw material also wraps
Methylene blue has been included, methylene blue powder is added after lecithin addition and has been stirred to dissolving, methylene blue concentration is 0.05- in the fixer of formation
0.15g/L。
The raw material further includes having PBS solution, and anhydrous methanol, acetone and formaldehyde are sequentially added in PBS solution and stirred equal
Even, PBS solution concentration is 100-200ml/L in the fixer of formation.
The raw material further includes having deoxycholic aicd, PBS solution, and anhydrous methanol, acetone and formaldehyde sequentially add in PBS solution
And after mixing evenly, lecithin and deoxycholic aicd are sequentially added, stirring is formed by fixer, deoxycholic aicd is dense to dissolving
Degree is 30~70g/L, and PBS solution concentration is 100-200ml/L.
The raw material further includes having deoxycholic aicd, methanol, PBS solution, and anhydrous methanol, methanol, acetone and formaldehyde successively add
Entering in PBS solution and after mixing evenly, sequentially adds lecithin and deoxycholic aicd, stirring is formed by fixer to dissolving,
Deoxycholic aicd concentration is 30~70g/L, and methanol concentration 200-300ml/L, PBS concentration is 100-200ml/L.
The raw material further includes having deoxycholic aicd, PBS solution and methylene blue, and anhydrous methanol, acetone and formaldehyde sequentially add PBS
In solution and after mixing evenly, lecithin, deoxycholic aicd and methylene blue are sequentially added, stirring is formed by fixer to dissolving,
Deoxycholic aicd concentration is 30~70g/L, and PBS solution concentration is 100-200ml/L, and methylene blue concentration is 0.05-0.15g/L.
The raw material further includes having deoxycholic aicd, methanol, PBS solution and methylene blue, anhydrous methanol, methanol, acetone and formaldehyde
It sequentially adds in PBS solution and after mixing evenly, sequentially adds lecithin, deoxycholic aicd and methylene blue, stirring is formed to dissolving
Fixer in, deoxycholic aicd concentration be 30~70g/L, methanol concentration 200-300ml/L, PBS solution concentration be 100-
200ml/L, methylene blue concentration are 0.05-0.15g/L.It is furthermore preferred that each component concentration is respectively as follows: formaldehyde in the fixer
100ml/L, lecithin 50g/L, deoxycholic aicd 47.5g/L, dehydrated alcohol 250ml/L, methanol 250ml/L, acetone 250ml/L,
PBS solution 1 ×, methylene blue 0.1g/L.
In process for preparation, first weighs dehydrated alcohol, acetone and formaldehyde (additionally increasing methanol, PBS solution when necessary) and stir
After mixing uniformly, it is added lecithin (increasing deoxycholic aicd, methylene blue powder when necessary), stirs to dissolve.
Above-mentioned palmitin fixer is handled into sample, advantage major embodiment are as follows:
(1) when the sample dissection being applied to after cancer routine radical surgery is with materials, in effective dissolution softening fat
While tissue, it is ensured that the timely abundant fixation of lymph node.
(2) the detection number of lymph node is improved, accurate lymphatic metastasis positive rate decreases the solution of Pathologis
The material time is taken, under the extremely short status of pathologist instantly, saves the more important and real meaning of human resources.
(3) possibility is provided for the outdoor quality control of later period pathology subject, in clinical wide popularization and application after commercialization
In effect it is especially prominent.
Detailed description of the invention
Fig. 1 is the adipose tissue of palmitin fixer before and after the processing prepared by the application, wherein A is the fresh rouge before processing
Fat tissue, B are fatty for treated；C is that metastatic carcinoma amplifies 40 times, and D is that small metastatic carcinoma amplifies 200 times；
Fig. 2 is breast tissue using palmitin fixer (uplink) prepared by the application and uses conventional neutral formalin
The effect comparison figure of (downlink) processing, wherein being followed successively by conventional H E dyeing and CD10, P63 and PR immunohistochemistry dye from left to right
Color, × 200；
Fig. 3 is thyroid gland sample using palmitin fixer (uplink) prepared by the application and uses conventional neutral formalin
The effect comparison figure of (downlink) processing, wherein being followed successively by conventional H E dyeing and TTF-1, CD20 and CD3 immunohistochemistry from left to right
Dyeing, × 200；
Fig. 4 is lung tissue sample using palmitin fixer (uplink) prepared by the application and uses conventional neutral formalin
The effect comparison figure of (downlink) processing, wherein being followed successively by conventional H E dyeing and CK, P63 and TTF-1 immunohistochemistry dye from left to right
Color, × 200；
Fig. 5 is colonic tissue sample using palmitin fixer (uplink) prepared by the application and uses conventional neutral formal
The effect comparison figure of woods (downlink) processing, wherein being followed successively by conventional H E dyeing and actin, CK20 and S100 immune group from left to right
Change and dyes, × 200.
Specific embodiment
Embodiment 1: differently composed palmitin fixer
The central role ingredient of palmitin fixer prepared by the application is mainly formaldehyde, lecithin, dehydrated alcohol and third
Ketone can also additionally increase any one of deoxycholic aicd, methanol, PBS solution and methylene blue or more under different implementation environments
Kind, specific ingredient selection mode can be found in shown in table 1.
The palmitin fixer of the different compositions of table 1
We respectively survey the palmitin fixer of core constituent (serial number 1 i.e. in table 1) and complementary element
Examination, available above-mentioned 16 kinds of situations, same composition do parallel test to same sample using same concentrations, the results showed that
The palmitin fixer of core constituent is whether to adipose tissue or organ-tissue (such as parathyroid tissue, breast tissue, lung
Tissue) or colonic tissue, all there is good fixed effect, lymph node is well exposed；But the effect of complementary element
Can not be ignored, as using serial number 2 in table 1 scheme is provided when, under the cooperation of deoxycholic aicd and core constituent, fatty disappears
It is preferable to change effect, the trend is in serial number 1 and serial number 2, serial number 3 and serial number 6, serial number 8 and serial number 12 and serial number 14 and serial number 16
Comparison in have same embodiment；In scheme provided by serial number 3, methanol plays the role of cooperateing with dehydrated alcohol, cooperates other
Core constituent, dissolution efficiency is higher, the trend in serial number 1 and serial number 3, serial number 4 and serial number 8, serial number 6 and serial number 7 and
There is same embodiment in the comparison of serial number 15 and serial number 16；In scheme provided by serial number 4, PBS solution and core constituent
Under cooperation, fatty dissolution and bating effect are more uniform, and the trend is in serial number 1 and serial number 4, serial number 13 and serial number 16；Serial number 5
In provided scheme, under the cooperation with core constituent, can more quickly it develop the color in subsequent dyeing processing, the spy
Property also have same embodiment in the comparison of serial number 1 and serial number 5, serial number 6 and serial number 13, serial number 12 and serial number 16.
Therefore, based on different fixed purpose and processing requirement, can choose above-mentioned differently composed palmitin fixer into
Row palmitin (fat melting) processing.
We also (remove core constituent in fixer (formaldehyde, lecithin, dehydrated alcohol, acetone) with complementary element
Oxycholic acid, methanol, PBS solution, methylene blue) concentration tested, the results showed that in palmitin fixer, the concentration of formaldehyde is suitable
The suitable control of concentration preferably controlled in 50-150ml/L, lecithin exists in the suitable control of concentration of 30-70g/L, dehydrated alcohol
200-300ml/L, acetone concentration be suitable for control in 200-300ml/L, in this case, four kinds of core action components can be with
It is good to play collaboration and mating reaction, achieve the effect that dissolution or softening fat, being lower than or exceed above range then can be right
Fat melting effect causes certain interference and adverse effect；When carrying out the addition of complementary element, above-mentioned four kinds of cores can be kept to make
With ingredient additive amount it is constant or within the above range appropriate adjustment, the addition of complementary element should not be too large, be otherwise easy to make
It is unbalance at fixer, softening and solute effect are influenced, therefore, concentration of the deoxycholic aicd in fixer is 30~70g/L, methanol
Concentration in fixer is 200-300ml/L, concentration of the PBS solution in fixer is 1 ×, methylene blue is dense in fixer
Degree is 0.05-0.15g/L, and these four complementary elements can add simultaneously, also may be selected any or appoints and several add respectively.
Embodiment 2: the palmitin fixer of same composition various concentration
This gives the full palmitins for constituting and (containing whole core constituent and whole complementary elements)
Fixer composition and concentration, and representative several groups are enumerated as shown in table 2:
The palmitin fixer of 2 various concentration of table
Pretreated operation is handled using palmitin fixer corresponding to each serial number of table 2 to cut off nethike embrane sample 4 hours, then
Lymph node materials, dehydration, embedding, slice, dyeing, immunohistochemistry are carried out, under the microscope, the results showed that above-mentioned each concentration
Different degrees of realizing to sample is dissolved, and the soft liquefaction of fat, lymph node is effectively exposed, and lymph node cells form has saved
Whole, no deformation, immunohistochemistry works well, when carrying out palmitin experiment especially with the provided scheme of serial number 9-12 in table 2,
Fat melting significant effect, good fixing effect, the soft liquefaction of fat, lymph node exposure is obvious, and lymph node cells form saves complete, nothing
Deformation, immunohistochemistry work well.
Embodiment 3: the effect comparison of different preparation methods
In our experimentation, it has been found that: each raw material (including central role ingredient as and complementary element such as go
Oxycholic acid, methanol, PBS solution, methylene blue) different orders of addition can also impact palmitin effect, specific manifestation are as follows: according to
Ethyl alcohol has the special type dissolved each other with a variety of organic solvents such as water methanol, ether with arbitrary proportion, first by methanol, acetone it is miscible with
In ethyl alcohol, then according to the hydrophily of lecithin, lecithin is dissolved in PBS (less than 50 degree in order to avoid failure), then in PBS
In successively add water-soluble formalin, be finally mixed and stirred for alcohol mixeding liquid and PBS mixed liquor uniformly.
Embodiment 4: application effect control
Individually below using palmitin fixer provided by serial number 12 in table 2 as representing, it is solid that palmitin is carried out to different specimens
Change, and is compared with conventional neutral formalin (10% formalin, pH7.2) treatment effect, specific as follows:
(1) adipose tissue
Operation excision fresh fat tissue fritter and lymph node is taken to be all made of same processing method in duplicate.First
With chemical Treatment, i.e., 10min is impregnated at room temperature with 0.01N HCl, treatment fluid is abandoned, after PBS is rinsed, with 0.25% tryptose
37 DEG C of constant water bath box of enzyme-EDTA impregnate 10min.In next step, by sample taking-up be put into degreasing fixer, then every
0.5-1h, which takes pictures, to be observed and recorded.As shown in Figure 1, this is former adipose tissue compared with degreasing fixer is handled after 2h, it is seen that
After being handled using the application palmitin fixer, adipose tissue obviously softens thinning, and lymph node structure is complete, metastatic carcinoma tissue and micro-
Small metastasis cancer cell group is high-visible.
(2) organ-tissue
It takes operation to cut off fresh mammary gland glandular tissue, first by chemical Treatment, i.e., is impregnated at room temperature with 0.01N HCl
10min, abandon treatment fluid PBS rinse after, then with 0.25% trypsase-EDTA in constant water bath box 37 DEG C of immersion 10min.Under
Sample taking-up is put into degreasing fixer by one step, and conventional materials, dehydration, paraffin embedding, slice and dye are successively carried out after 4h
Color, as a result as shown in Figure 2, the results showed that: fixed compared with sample with conventional formalin, the processing of this law fixer is to mammary gland
The equal indifference of general morphology, immunohistochemical staining of tissue specimen.
Take operation in cut off fresh thyroid gland glandular tissue, if upper type processing, as a result as shown in Figure 3, the results showed that: with it is normal
Advise that formalin is fixed compared with sample, general morphology of the processing of this law fixer to parathyroid tissue sample, immune group
Change and dyes equal indifference.
Operation is taken to cut off fresh lung tissue, if upper type is handled, as a result as shown in Figure 4, the results showed that: with conventional formal
Woods is fixed compared with sample.General morphology, immunohistochemical staining equal indifference of the processing of this law fixer to lung tissue sample
Not.
Operation excision colonic tissue is taken, if upper type is handled, as a result as shown in Figure 5, the results showed that: with conventional formalin
It fixes compared with sample.The processing of this law fixer is equal to general morphology, the immunohistochemical staining of colon mucosa tissues sample
Indifference.
Claims (10)
1. a kind of palmitin fixer, it is characterised in that: the fixer includes formaldehyde, lecithin, dehydrated alcohol and acetone, each group
The concentration in fixer is divided to be respectively as follows: 50 ~ 150ml/L of formaldehyde, 30 ~ 70g/L of lecithin, 200 ~ 300 ml/L of dehydrated alcohol,
200 ~ 300ml/L of acetone.
2. a kind of palmitin fixer as described in claim 1, it is characterised in that: further include having deoxycholic aicd, PBS solution and beauty
Orchid, concentration of the deoxycholic aicd in fixer are 30 ~ 70g/L, and it is 100-200ml/ that PBS solution, which is added to the concentration in fixer,
L, concentration of the methylene blue in fixer are 0.05-0.15g/L.
3. a kind of palmitin fixer as claimed in claim 2, which is characterized in that during the palmitin fixer is constituted, each component
Concentration is respectively as follows: formaldehyde 100ml/L, lecithin 50g/L, deoxycholic aicd 47.5g/L, 250 ml/L of dehydrated alcohol, methanol 250
Ml/L, acetone 250ml/L, PBS solution 1 ×.
4. a kind of palmitin fixer as described in claim 1, it is characterised in that: further include having deoxycholic aicd, methanol, PBS molten
Liquid and methylene blue, concentration of the deoxycholic aicd in fixer are 30 ~ 70g/L, and concentration of the methanol in fixer is 200-300ml/
L, it is 100-200ml/L that PBS solution, which is added to the concentration in fixer, and concentration of the methylene blue in fixer is 0.05-0.15g/
L。
5. a kind of palmitin fixer as claimed in claim 4, which is characterized in that during the palmitin fixer is constituted, each component
Concentration is respectively as follows: formaldehyde 100ml/L, lecithin 50g/L, deoxycholic aicd 47.5g/L, 250 ml/L of dehydrated alcohol, methanol 250
Ml/L, acetone 250ml/L, PBS solution 1 ×, 0. 1g/L of methylene blue.
6. a kind of preparation method of palmitin fixer, it is characterised in that: using formaldehyde, lecithin, dehydrated alcohol and acetone as original
Material, after mixing evenly by dehydrated alcohol, formaldehyde and acetone, be added lecithin, stir and make it dissolve, in the fixer of formation on
The concentration for stating each raw material is respectively as follows: 50 ~ 150ml/L of formaldehyde, 30 ~ 70g/L of lecithin, 200 ~ 300 ml/L of dehydrated alcohol, acetone
200~300ml/L。
7. a kind of preparation method of palmitin fixer as claimed in claim 6, it is characterised in that: the raw material further includes having
Oxycholic acid is added deoxycholic aicd and stirs to dissolving after lecithin addition, in the fixer of formation deoxycholic aicd concentration be 30 ~
70g/L。
8. a kind of preparation method of palmitin fixer as claimed in claim 6, it is characterised in that: the raw material further includes having first
Alcohol, methanol concentration is 200-300ml/L in the fixer of formation.
9. a kind of preparation method of palmitin fixer as claimed in claim 6, it is characterised in that: the raw material further includes having beauty
Orchid is added methylene blue powder and stirs to dissolving after lecithin addition, methylene blue concentration is 0.05-0.15g/L in the fixer of formation.
10. a kind of preparation method of palmitin fixer as claimed in claim 6, it is characterised in that: the raw material further includes having
PBS solution, anhydrous methanol, acetone and formaldehyde are sequentially added in PBS solution and are stirred evenly, PBS solution in the fixer of formation
Concentration is 100-200ml/L.
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