CN101182506A - Fixation fluid for saving cast-off cells and manufacturing method of cell shallow layer smear - Google Patents
Fixation fluid for saving cast-off cells and manufacturing method of cell shallow layer smear Download PDFInfo
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- CN101182506A CN101182506A CNA2007100504639A CN200710050463A CN101182506A CN 101182506 A CN101182506 A CN 101182506A CN A2007100504639 A CNA2007100504639 A CN A2007100504639A CN 200710050463 A CN200710050463 A CN 200710050463A CN 101182506 A CN101182506 A CN 101182506A
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Abstract
The invention discloses a preservation solution for exfoliated cell and a preparation method which uses the preservation solution to process cell thin layer smear. The preservation solution for exfoliated cell comprises the components which are prepared by the volume proportion of 10 to 15 portions of methanol, 10 to 15 portions of alcohol, 1 to 2 portions of glycerin, 1 to 2 portions of glycol polyethylene, 10 to 12 portions of formaldehyde, 0.5 to 1 portions of glacial acetic acid, 10 to 20 portions of sodium chloride with 0.85 percent to 0.9 percent of mass concentration and 33 percent to 68.5 percent of phosphate buffer. The steps of the preparation method for cell thin layer smear are that: 1) the collected sample is put in the preservation solution for exfoliated cell for shaking; the sample is transferred to a centrifuge for centrifugal treatment after standing; 2) the supernatant is removed until 1ml to 2ml residue is left in the centrifuge tube; the cell suspension is obtained after uniform vibration; 3) 0.5ml to 1ml cell suspension is put into a smear preparation cup which is made into cell thin layer smear by a cell smear preparation machine.
Description
(1) technical field:
The present invention relates to a kind of cell and preserve stationary liquid, be specifically related to cast-off cells and preserve stationary liquid; The invention still further relates to the making method of the cell thin smear that uses this preservation stationary liquid processing.
(2) background technology:
Viable cell has physiological function, biologic activity and inherent cellular form and structure, has using value aspect a lot.But, if being broken away from original living environment, it places, and the form of cells physiological function and biologic activity and cell and structure will change, and ordinary method is that pair cell carries out freezing preservation.But freezing preservation need be bought special refrigerating apparatus, and, owing to freezing, the operation of thawing will inevitably produce significant infringement by pair cell, thereby cause the variation distortion of cellular form, structure.Therefore, for the routine inspection of cast-off cells, freezing preservation is not the best way.
It is reported that if cell suspending liquid is placed in the solution that contains pure and mild sequestrant, cell can be preserved about 3 week under about 37 ℃ of conditions.
Preservation liquid (the PresetverCyt of the present new Bai Shi of the U.S. (Tinpreper TCT) manufacturer production
TMSlution) be and the matching used reagent of its machine Tinpreper-2000, but costing an arm and a leg of it, in order to reduce cost, the supporting cell-preservation liquid of many and TCT arises at the historic moment, as application number is 200610017591.9, name is called the Chinese invention patent of " exfoliated cell preservative fluid ", disclose a kind of exfoliated cell preservative fluid, it is grouped into by the following one-tenth of part meter by volume: 40～60 parts of methyl alcohol or ethanol, 20～40 parts of phosphate buffered saline buffer or Tutofusin tris hydrochloride buffers, 5～10 parts in formaldehyde or acetone, 5～10 parts of EDTA-2K or disodium ethylene diamine tetraacetate.The starting material of this exfoliated cell preservative fluid are common biochemical reagents, thereby have reduced the price of TCT consumptive material.Secondly can reduce and go out the dissatisfied rate of blood specimen for the first time, reduce the consumption and the waste of time of the consumptive material of handling sample again and being brought; Because penetration power forces cell fixation abundant, keep cell fixing under virgin state as far as possible.
Existing making cell thin smear has two kinds of technology: the one, and liquid based thin-layer cell technology (TCT technology), it is that the cell that will obtain adds in the preservation liquid, directly on sheet glass, make the thin-layer cell smear by the method that the machine automatic negative-pressure attracts, shifts, its advantage is the automatic film-making of machine, shortcoming is to make long, machine expense height of a thin slice and film-making time, maintenance complexity etc. at every turn; The 2nd, the LPT technology is cell suspension directly to be dropped in be made into smear on the sheet glass, and its advantage is to need not specific installation, and is simple to operate, and its shortcoming is the cell that obtains all not to be used to make smear, causes cell overlap easily, is unfavorable for inspections and examinations.
(3) summary of the invention:
The present invention will disclose that a kind of cellular segregation is effective, impurity solubleness height, cell curing is effective and cell bonding die rate is high cast-off cells are preserved stationary liquid, the making method of the cell thin smear that the present invention also handles this preservation stationary liquid of public use.
Cast-off cells of the present invention are preserved stationary liquid, and it comprises the component of following proportioning in parts by volume:
Methyl alcohol 10-15 part; Ethanol 10-15 part; Glycerol 1-2 part;
Polyoxyethylene glycol 1-2 part; Formaldehyde 10-12 part; Glacial acetic acid 0.5-1 part;
Mass concentration is sodium-chlor 10-20 part of 0.85-0.9%;
Phosphoric acid buffer 33-68.5 part.
Described methyl alcohol, ethanol, glycerol, polyoxyethylene glycol, formaldehyde, Glacial acetic acid and sodium-chlor are at least chemical pure reagent; Wherein, the volumetric concentration of methyl alcohol is 〉=99%, and the alcoholic acid volumetric concentration is 〉=99%, and the volumetric concentration of glycerol is 〉=99%, and the molecular weight of polyoxyethylene glycol can be 180-220, and the volumetric concentration of formaldehyde can be 37-40%, and the volumetric concentration of Glacial acetic acid is 〉=99%; The purity of described sodium-chlor 〉=99%;
The pH value of described phosphoric acid buffer is preferably 6.3-6.5.
The present invention also comprises the making method of cell thin smear, and step is as follows:
1) sample of gathering placed cast-off cells preserve the stationary liquid vibration, changes whizzer over to after leaving standstill and carry out centrifugal treating;
2) pour out supernatant liquid, the residuum in centrifuge tube is 1-2ml, it is shaken up again, and obtains cell suspending liquid;
3) get the 0.5-1ml cell suspending liquid and put into the film-making cup, make cell thin smear with the cell pelleter;
Wherein, described exfoliated cell preservative fluid comprises the component of following proportioning in parts by volume:
Methyl alcohol 10-15 part; Ethanol 10-15 part; Glycerol 1-2 part;
Polyoxyethylene glycol 1-2 part; Formaldehyde 10-12 part; Glacial acetic acid 0.5-1 part;
Mass concentration is sodium-chlor 10-20 part of 0.85-0.9%;
Phosphoric acid buffer 33-68.5 part.
Described methyl alcohol, ethanol, glycerol, polyoxyethylene glycol, formaldehyde, Glacial acetic acid and sodium-chlor are at least chemical pure reagent; Wherein, the volumetric concentration of methyl alcohol is 〉=99%, and the alcoholic acid volumetric concentration is 〉=99%, and the volumetric concentration of glycerol is 〉=99%, and the molecular weight of polyoxyethylene glycol can be 180-220, and the volumetric concentration of formaldehyde can be 37-40%, and the volumetric concentration of Glacial acetic acid is 〉=99%; The purity of described sodium-chlor 〉=99%;
The pH value of described phosphoric acid buffer is preferably 6.3-6.5;
In the above-mentioned making method step 1), the time of described vibration is preferably 10-20 minute, so that cell separates fully; The described time of leaving standstill is preferably 10-30 minute, so that cell is fully fixing, makes the impurity dissolving simultaneously fully; Rotating speed during described centrifugal treating can be 2000 rev/mins, and the time is preferably 5-8 minute; In the step 3), the cell filtration device in the described cell pelleter is stained with 100-200 purpose filter screen for the hollow space at filter paper or thieving paper.
What cast-off cells of the present invention were preserved the stationary liquid use all is general chemistry reagent, thereby has reduced the price of TCT consumptive material; It can dissolve red corpuscle, cell debris and little viscose rayon more than 95%, has improved the separating effect of cell, thereby has improved the production effect of postorder, makes that thin-layer cell sheet background is clear, impurity is few, and cell distribution is even; In addition, owing in preserving liquid, added formaldehyde, thus improved the solidification effect of cell greatly; The cytomorphosis rate is low, can keep original form to stick on the slide glass.
The making method of cell thin smear of the present invention is handled cast-off cells owing to adopted above-mentioned cast-off cells to preserve stationary liquid, and the adhesion rate of cell is high and be difficult for flake; And in the film-making process, adopted hollow space to be stained with the filter paper or the thieving paper of filter screen, and not only removed a large amount of impurity, also make the thin smear cell distribution of making even, help inspections and examinations, thereby improved positive rate greatly.
(4) embodiment:
Embodiment 1: cell is preserved the preparation of stationary liquid
Below used reagent be chemical pure.
Getting the 10ml volumetric concentration and be 99% methyl alcohol, 15ml volumetric concentration and be 100% ethanol, 2ml volumetric concentration and be 99% glycerol, 2ml molecular weight and be 180 polyoxyethylene glycol, 10ml volumetric concentration and be 37% formaldehyde, 0.5ml volumetric concentration and be 100% Glacial acetic acid, 10ml mass concentration and be 0.9% sodium-chlor and 68.5ml pH value and be 6.3 phosphoric acid buffer mixes and promptly obtains cast-off cells of the present invention and preserve stationary liquid.
| The preservation liquid of Tinpreper TCT manufacturer production | Embodiment 1 described cast-off cells are preserved stationary liquid | |
| Price | 60 yuan/person-portion | 45 yuan/person-portion |
| The sample floss | Many | Few |
| The cell aggregation amount | Many | Few |
Embodiment 2: cell is preserved the preparation of stationary liquid
Below used reagent be analytical pure.
Getting the 15ml volumetric concentration and be 99% methyl alcohol, 15ml volumetric concentration and be 99% ethanol, 1ml volumetric concentration and be 99% glycerol, 2ml molecular weight and be 200 polyoxyethylene glycol, 12ml volumetric concentration and be 40% formaldehyde, 1ml volumetric concentration and be 99% Glacial acetic acid, 20ml mass concentration and be 0.85% sodium-chlor and 34ml pH value and be 6.5 phosphoric acid buffer mixes and promptly obtains cast-off cells of the present invention and preserve stationary liquid.
Embodiment 3: the making method of cell thin smear
1) sample of gathering is placed 10ml embodiment 2 described cast-off cells preserve stationary liquids vibration 20 minutes, leave standstill again pour into after 10 minutes in the centrifuge tube with 2000 rev/mins 8 minutes;
2) pour out supernatant liquid, the residuum in centrifuge tube is 1-2ml, it is shaken up again, and obtains cell suspending liquid;
3) get the 0.5ml cell suspending liquid and put into the film-making cup,, make cell thin smear with containing the cell pelleter that hollow space is stained with the filter paper of 100 mesh filter screens.
Claims (10)
1. cast-off cells are preserved stationary liquid, and it is characterized in that: it comprises the component of following proportioning in parts by volume:
Methyl alcohol 10-15 part; Ethanol 10-15 part; Glycerol 1-2 part;
Polyoxyethylene glycol 1-2 part; Formaldehyde 10-12 part; Glacial acetic acid 0.5-1 part;
Mass concentration is sodium-chlor 10-20 part of 0.85-0.9%;
Phosphoric acid buffer 33-68.5 part.
2. cast-off cells according to claim 1 are preserved stationary liquid, it is characterized in that: the pH value of described phosphoric acid buffer is 6.3-6.5.
3. the making method of a cell thin smear, step is as follows:
1) sample of gathering placed cast-off cells preserve the stationary liquid vibration, changes whizzer over to after leaving standstill and carry out centrifugal treating;
2) pour out supernatant liquid, the residuum in centrifuge tube is 1-2ml, it is shaken up again, and obtains cell suspending liquid;
3) get the 0.5-1ml cell suspending liquid and put into the film-making cup, make cell thin smear with the cell pelleter;
Wherein, described exfoliated cell preservative fluid comprises the component of following proportioning in parts by volume:
Methyl alcohol 10-15 part; Ethanol 10-15 part; Glycerol 1-2 part;
Polyoxyethylene glycol 1-2 part; Formaldehyde 10-12 part; Glacial acetic acid 0.5-1 part;
Mass concentration is sodium-chlor 10-20 part of 0.85-0.9%;
Phosphoric acid buffer 33-68.5 part.
4. the making method of cell thin smear according to claim 3 is characterized in that: the pH value of described phosphoric acid buffer is 6.3-6.5.
5. according to the making method of claim 3 or 4 described cell thin smears, it is characterized in that: in the step 3), described cell pelleter is provided with the cell filtration device.
6. the making method of cell thin smear according to claim 5 is characterized in that: described cell filtration device is stained with filter screen for the hollow space at filter paper or thieving paper.
7. the making method of cell thin smear according to claim 6, it is characterized in that: described filter screen is the 100-200 order.
8. according to the making method of claim 3 or 4 described cell thin smears, it is characterized in that: in the step 1), the time of described vibration is 10-20 minute.
9. according to the making method of claim 3 or 4 described cell thin smears, it is characterized in that: in the step 1), the described time of leaving standstill is 10-30 minute.
10. according to the making method of claim 3 or 4 described cell thin smears, it is characterized in that: in the step 1), the rotating speed during described centrifugal treating is 2000 rev/mins, and the time is 5-8 minute.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2007100504639A CN101182506A (en) | 2007-11-07 | 2007-11-07 | Fixation fluid for saving cast-off cells and manufacturing method of cell shallow layer smear |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2007100504639A CN101182506A (en) | 2007-11-07 | 2007-11-07 | Fixation fluid for saving cast-off cells and manufacturing method of cell shallow layer smear |
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| CN101182506A true CN101182506A (en) | 2008-05-21 |
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| CNA2007100504639A Pending CN101182506A (en) | 2007-11-07 | 2007-11-07 | Fixation fluid for saving cast-off cells and manufacturing method of cell shallow layer smear |
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101999343A (en) * | 2010-10-26 | 2011-04-06 | 深圳华大基因科技有限公司 | Cell preserving fluid and preparation method and use thereof |
| CN103461320A (en) * | 2013-08-19 | 2013-12-25 | 安徽信灵检验医学科技有限公司 | Non-isotonic preserving fluid for liquid-based thin-layer exfoliative cells |
| CN104041484A (en) * | 2014-05-23 | 2014-09-17 | 科蒂亚(新乡)生物技术有限公司 | Cell preservation liquid |
| CN104642300A (en) * | 2015-03-13 | 2015-05-27 | 上海御康医院有限公司 | Exfoliative cell sap base preserving fluid, method thereof for flaking and kit |
| CN105807044A (en) * | 2016-02-04 | 2016-07-27 | 山西瑞豪生物科技有限公司 | Method for fixing target-antigen cells of inflammatory bowel disease |
| CN105994252A (en) * | 2016-06-22 | 2016-10-12 | 杭州海世嘉生物科技有限公司 | Liquid-based thin cell preservation liquid and preparation method thereof |
| CN107367417A (en) * | 2017-07-14 | 2017-11-21 | 上海市松江区中心医院 | A kind of quick fixative composition of Ultrasound-guided Biopsy cell and preparation method thereof |
| CN110786318A (en) * | 2019-11-12 | 2020-02-14 | 杭州昱鼎生物科技有限公司 | Urine cell pre-fixing liquid |
| CN111670896A (en) * | 2020-04-28 | 2020-09-18 | 浙江大学医学院附属妇产科医院 | Neutral buffer tissue fixing solution |
| CN111972396A (en) * | 2019-05-21 | 2020-11-24 | 威海威高医用材料有限公司 | Cervical exfoliated cell preservation solution, preparation method and cell preservation method |
-
2007
- 2007-11-07 CN CNA2007100504639A patent/CN101182506A/en active Pending
Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101999343A (en) * | 2010-10-26 | 2011-04-06 | 深圳华大基因科技有限公司 | Cell preserving fluid and preparation method and use thereof |
| CN101999343B (en) * | 2010-10-26 | 2013-11-06 | 深圳华大基因健康科技有限公司 | Cell preserving fluid and preparation method and use thereof |
| CN103461320A (en) * | 2013-08-19 | 2013-12-25 | 安徽信灵检验医学科技有限公司 | Non-isotonic preserving fluid for liquid-based thin-layer exfoliative cells |
| CN103461320B (en) * | 2013-08-19 | 2015-09-02 | 安徽信灵检验医学科技有限公司 | An a kind of conserving liquid such as not grade for liquid base thin layer cast-off cells |
| CN104041484A (en) * | 2014-05-23 | 2014-09-17 | 科蒂亚(新乡)生物技术有限公司 | Cell preservation liquid |
| CN104642300A (en) * | 2015-03-13 | 2015-05-27 | 上海御康医院有限公司 | Exfoliative cell sap base preserving fluid, method thereof for flaking and kit |
| CN105807044A (en) * | 2016-02-04 | 2016-07-27 | 山西瑞豪生物科技有限公司 | Method for fixing target-antigen cells of inflammatory bowel disease |
| CN105994252A (en) * | 2016-06-22 | 2016-10-12 | 杭州海世嘉生物科技有限公司 | Liquid-based thin cell preservation liquid and preparation method thereof |
| CN105994252B (en) * | 2016-06-22 | 2019-07-02 | 杭州海世嘉生物科技有限公司 | A kind of Thinprep pap test saves liquid and preparation method thereof |
| CN107367417A (en) * | 2017-07-14 | 2017-11-21 | 上海市松江区中心医院 | A kind of quick fixative composition of Ultrasound-guided Biopsy cell and preparation method thereof |
| CN107367417B (en) * | 2017-07-14 | 2021-06-22 | 上海市松江区中心医院 | Rapid fixing agent composition for punctured cells under ultrasonic guidance and preparation method thereof |
| CN111972396A (en) * | 2019-05-21 | 2020-11-24 | 威海威高医用材料有限公司 | Cervical exfoliated cell preservation solution, preparation method and cell preservation method |
| CN110786318A (en) * | 2019-11-12 | 2020-02-14 | 杭州昱鼎生物科技有限公司 | Urine cell pre-fixing liquid |
| CN111670896A (en) * | 2020-04-28 | 2020-09-18 | 浙江大学医学院附属妇产科医院 | Neutral buffer tissue fixing solution |
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Open date: 20080521 |