WO2019161653A1 - Method for staining pathological sections - Google Patents
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- WO2019161653A1 WO2019161653A1 PCT/CN2018/103399 CN2018103399W WO2019161653A1 WO 2019161653 A1 WO2019161653 A1 WO 2019161653A1 CN 2018103399 W CN2018103399 W CN 2018103399W WO 2019161653 A1 WO2019161653 A1 WO 2019161653A1
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- 230000001575 pathological effect Effects 0.000 title claims abstract description 35
- 238000000034 method Methods 0.000 title claims abstract description 31
- 238000010186 staining Methods 0.000 title claims abstract description 28
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 70
- 238000004043 dyeing Methods 0.000 claims description 18
- 239000007788 liquid Substances 0.000 claims description 7
- 241001510071 Pyrrhocoridae Species 0.000 claims description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 3
- 229910052799 carbon Inorganic materials 0.000 claims description 3
- 229930195733 hydrocarbon Natural products 0.000 claims description 3
- 239000004215 Carbon black (E152) Substances 0.000 claims description 2
- 125000001183 hydrocarbyl group Chemical group 0.000 claims description 2
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 150000001875 compounds Chemical class 0.000 claims 1
- 238000011534 incubation Methods 0.000 abstract description 9
- 238000007447 staining method Methods 0.000 description 8
- 230000000694 effects Effects 0.000 description 5
- 238000011532 immunohistochemical staining Methods 0.000 description 3
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Chemical compound C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- YQGOJNYOYNNSMM-UHFFFAOYSA-N eosin Chemical compound [Na+].OC(=O)C1=CC=CC=C1C1=C2C=C(Br)C(=O)C(Br)=C2OC2=C(Br)C(O)=C(Br)C=C21 YQGOJNYOYNNSMM-UHFFFAOYSA-N 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 150000002430 hydrocarbons Chemical class 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 238000010339 medical test Methods 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
Classifications
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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
Abstract
Disclosed is a method for staining pathological sections, which is used for an automated staining system and comprises the steps of adding, during a staining process, reaction reagents and leaving same to stand for incubation for 1-10 minutes before mixing the reaction reagents homogeneously. The technical solution improves methods for staining pathological sections, involves the homogeneous mixing of reaction reagents during an incubation process, improves the use efficiency of the reagents, and saves on the usage amount of the reagents.
Description
一种病理切片的染色方法 Method for staining pathological sections
[0001] 本申请是以申请号为 201810159656.6、 申请日为 2018年 2月 26日的中国专利申请 为基础， 并主张其优先权， 该申请的全部内容在此作为整体引入本申请中。 [0001] The present application is based on a Chinese patent application filed on Jan. 26, 2011, the entire disclosure of which is hereby incorporated by reference.
[0002] 技术领域 Technical Field
[0003] 本申请涉及医学检验的技术领域， 尤其涉及一种病理组织切片的染色方法。 [0003] The present application relates to the technical field of medical examination, and in particular to a method for staining a pathological tissue section.
[0004] 背景技术 BACKGROUND OF THE INVENTION
[0005] 病理切片染色是医学检验中的重要手段， 当前的染色方法包括常规染色、 IHC 染色、 特殊染色等， 需要用到各种试剂。 5见有染色方法中， 加入反应试剂后孵 育的过程中， 一般会保持静置载玻片， 这样造成离组织较远的反应试剂不能充 分参与反应， 造成试剂的浪费。 因而如果能使离组织较远的反应试剂也参与免 疫反应， 将提高试剂的使用效率。 5见有自动化染色方案， 染色时为了防止试剂 挥发， 有时会在载玻片上增加盖子或者添加液态油膜。 载破片和盖子放置在染 色仪特定的托盘上， 盖子除了能够防止挥发还能够保持组织上试剂的温度稳定 ， 增强染色效果。 同理地， 液态油膜也起到了防止挥发的作用。 但是如前所述 ， 为了使组织充分被试剂覆盖， 染色时通常需要将整个载玻片都覆盖上试剂， 大量的试剂不会参与反应， 造成不必要的浪费。 如果能使所有试剂都参与反应 ， 将能大大节省试剂用量。 [0005] Pathological section staining is an important means in medical tests, and current staining methods include conventional staining, IHC staining, special staining, etc., and various reagents are required. 5 In the dyeing method, during the incubation process after adding the reaction reagent, the slide glass is generally left to stand still, so that the reaction reagent far away from the tissue cannot fully participate in the reaction, resulting in waste of the reagent. Therefore, if the reaction reagent farther from the tissue can be involved in the immunological reaction, the use efficiency of the reagent will be improved. 5 See automatic dyeing scheme. In order to prevent the reagent from volatilizing during dyeing, sometimes cover or add liquid oil film on the slide. The fragment and the cover are placed on a specific tray of the dye meter. In addition to preventing evaporation, the lid can keep the temperature of the reagent on the tissue stable and enhance the dyeing effect. Similarly, the liquid oil film also acts to prevent volatilization. However, as mentioned above, in order to make the tissue fully covered by the reagent, it is usually necessary to cover the entire slide with the reagent, and a large amount of the reagent does not participate in the reaction, resulting in unnecessary waste. If all the reagents are involved in the reaction, the amount of the reagent can be greatly saved.
[0006] 申请内容 [0006] Application Content
[0007] 本申请的目的在于提供一种病理组织切片的染色方法， 减少了染色时试剂的使 用量， 提高试剂的使用效率。 为了实现上述目的， 本申请提供如下技术方案： 一种病理切片的染色方法， 应用于自动化染色系统， 包括步骤： [0007] The purpose of the present application is to provide a method for staining a pathological tissue section, which reduces the amount of reagent used in dyeing and improves the efficiency of use of the reagent. In order to achieve the above object, the present application provides the following technical solutions: A staining method for pathological sections, applied to an automated dyeing system, including steps:
[0008] b、 向病理切片加入反应试剂； [0008] b, adding a reaction reagent to the pathological section;
[0009] c、 静置孵育 1-10分钟； [0009] c, static incubation for 1-10 minutes;
[0010] d、 完成步骤 c后， 对反应试剂进行混匀； [0010] d, after the completion of step c, the reaction reagent is mixed;
[0011] 优选地， 在步骤 d之后还包括步骤： e、 完成步骤 d之后，至少重复一次步骤 c、 步骤 d操作。
[0012] 优选地， 染色过程中使用载玻片盖体； 所述载玻片盖体包括反应腔、 加样槽和 突起部； 载玻片盖体放置于自动化染色仪托盘内， 盖体的反应腔位置对应载玻 片上的组织切片位置。 [0011] Preferably, after step d, the method further comprises the steps of: e. After the step d is completed, the steps c and d are repeated at least once. [0012] Preferably, the slide cover is used in the dyeing process; the slide cover comprises a reaction chamber, a sample loading slot and a protrusion; the slide cover is placed in the automatic stainer tray, and the cover is The position of the reaction chamber corresponds to the position of the tissue slice on the slide.
[0013] 优选地， 所述步骤 b具体包括： 在盖体加样槽加入反应试剂。 [0013] Preferably, the step b specifically includes: adding a reaction reagent in the cover sample loading tank.
[0014] 优选地， 所述步骤 d具体包括： 将盖体突出部抬起再放回原来位置， 静置载玻 片后， 重复操作， 使得反应试剂充分混匀。 [0014] Preferably, the step d specifically includes: lifting the lid protrusion and returning to the original position, and after the glass slide is left, repeating the operation, so that the reaction reagent is sufficiently mixed.
[0015] 优选地， 所述步骤 d具体包括： 将盖体突出部抬起并向突出部方向拖动后再将 放置回原来位置， 静置载玻片后， 重复操作， 使得反应试剂充分混匀。 [0015] Preferably, the step d specifically includes: lifting the cover protrusion and dragging it in the direction of the protrusion, and then returning to the original position, after the slide is left, the operation is repeated, so that the reaction reagent is thoroughly mixed. uniform.
[0016] 优选地， 染色过程中使用液态油膜； 所述油膜包裹于病理切片外； 油膜成分为 含碳数为 C10-C16的碳氢化合物。 [0016] Preferably, a liquid oil film is used in the dyeing process; the oil film is wrapped outside the pathological section; and the oil film component is a hydrocarbon having a carbon number of C10-C16.
[0017] 优选地， 所述步骤 b具体包括： 向病理切片加入试剂， 试剂穿过油膜与病理切 片接触， 油膜包裹于反应试剂外。 [0017] Preferably, the step b specifically comprises: adding a reagent to the pathological section, the reagent is in contact with the pathological slice through the oil film, and the oil film is wrapped outside the reaction reagent.
[0018] 优选地， 所述步骤 d具体包括： 向由油膜包裹的反应试剂提供气流， 气流的作 用下反应试剂在油膜范围内流动； 静置后， 重复操作， 使得反应试剂充分混匀 [0018] Preferably, the step d specifically includes: providing a gas flow to the reaction reagent wrapped by the oil film, and flowing the reaction reagent in the oil film range under the action of the gas flow; after standing, repeating the operation to fully mix the reaction reagent
[0019] 优选地， 所述油膜成分为 C 15H 32。 [0019] Preferably, the oil film component is C 15 H 32 .
[0020] 本申请的技术方案改进了病理切片染色方法， 在孵育过程中混匀反应试剂， 提 高试剂的使用效率， 节约试剂的总用量。 [0020] The technical solution of the present application improves the pathological section staining method, and mixes the reaction reagent during the incubation process, thereby improving the use efficiency of the reagent and saving the total amount of the reagent.
[0021] 具体实施方式 DETAILED DESCRIPTION
[0022] 为使技术人员更好地理解本申请， 下面对本申请的实施例进行清楚、 详细的说 明， 但不作为对本申请的限定。 显然， 所描述的实施例仅仅是本申请的一部分 实施例， 而不是全部的实施例。 基于本申请中的实施例， 本领域普通技术人员 在没有作出创造性劳动前提下所获得的所有其他实施例， 都属于本申请保护的 范围。 [0022] The embodiments of the present application are clearly and in detail described below, but are not intended to limit the invention. It is apparent that the described embodiments are only a part of the embodiments of the present application, and not all of them. All other embodiments obtained by a person of ordinary skill in the art based on the embodiments of the present application without departing from the inventive scope are the scope of the present application.
[0023] 在一具体实施例中， 本申请公开的病理组织切片染色方法， 包括如下步骤： [0023] In a specific embodiment, the method for staining a pathological tissue section disclosed in the present application includes the following steps:
[0024] ( 1) 将贴有病理切片的载玻片放置在自动化染色仪托盘内； [0024] (1) placing the slide with the pathological section placed in an automated stainer tray;
[0025] (2) 将载玻片盖体放置于载玻片上， 盖体的反应腔位置对应载玻片上的组 织切片位置;
[0026] (3) 在盖体加样槽加入反应试剂； [0025] (2) placing the slide cover on the slide, the position of the reaction chamber of the cover corresponds to the position of the tissue slice on the slide; [0026] (3) adding a reagent in the lid sample loading tank;
[0027] (4) 混匀反应试剂： 试剂孵育 1-10分钟后， 将盖体突出部抬起再放回原来 位置， 或者将盖体突出部抬起向突出部方向拖动后再将放置回原来位置； [0027] (4) Mixing the reaction reagent: After the reagent is incubated for 1-10 minutes, lift the lid protrusion and put it back to the original position, or lift the lid protrusion to the direction of the protrusion and then place it. Return to the original position;
[0028] (5) 静置载玻片后， 重复步骤 (4) 操作， 使得反应试剂充分混匀。 上述重 复混匀的步骤， 使得原本远离组织、 未参与反应的试剂移动到组 [0028] (5) After standing the slide, repeat step (4) to allow the reaction reagent to mix well. The above repeated mixing step moves the reagents that are originally away from the tissue and are not involved in the reaction to the group
[0029] 织上方， 与组织进行反应， 提高试剂的使用效率， 在达到同样效果的前提下， 节省试剂的用量。 以 Ki67染色为例， 5见有的染色方法需要的抗体试剂用量约 [0030] lOOul； 应用本申请的载玻片盖体和上述染色方法进行 Ki67染色时， 达到同样 染色效果所需抗体试剂用量仅为 70ul。 同理的， 在其它多种染色中， 应用上述 方法后试剂使用量可减少 20以上， 大大节约了染色成本。 [0029] Above the weave, reacting with the tissue to increase the efficiency of use of the reagent, and saving the amount of the reagent while achieving the same effect. Take Ki67 staining as an example, 5 see the amount of antibody reagent required for some staining methods [0030] lOOul; the amount of antibody reagent required to achieve the same staining effect when using the slide cover of the present application and the above staining method for Ki67 staining Only 70ul. Similarly, in other kinds of dyeing, the amount of reagents can be reduced by more than 20 after applying the above method, and the dyeing cost is greatly saved.
[0031] 在一具体实施例中， 本申请公开的病理切片染色方法， 包括如下步骤： [0031] In a specific embodiment, the pathological section staining method disclosed in the present application includes the following steps:
[0032] (a) 将贴有病理切片的载玻片放置在自动化染色仪托盘内； [0032] (a) placing the slide with the pathological section placed in an automated stainer tray;
[0033] ⑹ 向病理组织切片位置滴加成分为 C 15H 32的液态油膜； [0033] (6) adding a liquid oil film having a composition of C 15 H 32 to the pathological tissue section;
[0034] (c) 向病理切片加入试剂， 试剂穿过油膜与病理切片接触， 油膜包裹于试 剂外； [0034] (c) adding a reagent to the pathological section, the reagent is in contact with the pathological section through the oil film, and the oil film is wrapped outside the reagent;
[0035] (d) 混匀反应试剂： 试剂孵育 1-10分钟后， 向由油膜包裹的反应试剂提供 气流， 气流的作用使得反应试剂在油膜范围内流动； 或者通过摇晃载玻片， 使 得反应试剂可流动， 已达到未反应的试剂流动到病理切片上方参与反应的效果 [0035] (d) Mixing the reaction reagent: After the reagent is incubated for 1-10 minutes, a gas stream is supplied to the reaction reagent wrapped by the oil film, and the gas stream acts to cause the reagent to flow in the oil film range; or by shaking the slide to make the reaction The reagent can flow, and the unreacted reagent has flowed to the top of the pathological section to participate in the reaction.
[0036] (e) 静置后， 重复上述步骤 (d) 操作 1-3次， 使得反应试剂充分混匀。 上 述使用液态油膜的染色方法， 在孵育过程中对反应试剂进行混匀， 使 [0036] (e) After standing, repeat the above step (d) operation 1-3 times to allow the reaction reagent to be thoroughly mixed. The above method of dyeing using a liquid oil film, mixing the reaction reagent during the incubation, so that
[0037] 得原反应的试剂最大限度得参与反应， 提高试剂的使用效率， 在达到同样效果 的前提下， 节省试剂的用量。 上述染色方法所应用的液态油膜还可以是其他含 碳数为 C10-C16的碳氢化合物。 [0037] The reagent for obtaining the original reaction is maximally involved in the reaction, and the use efficiency of the reagent is improved, and the amount of the reagent is saved under the premise of achieving the same effect. The liquid oil film to which the above dyeing method is applied may also be other hydrocarbons having a carbon number of C10-C16.
[0038] 本申请的染色方法中， 具体孵育时间根据具体反应试剂而定。 本申请的染色方 法应用于免疫组化染色、 常规染色和特殊染色。 在免疫组化染色中， 一抗孵育 、 二抗孵育、 DAB染色等均可应用本技术方案所公开的混匀反应试剂的操作方 法； 同样地， 本申请的技术方案可应用于在伊红染色、 苏木素染色等常规染色
中。 [0038] In the staining method of the present application, the specific incubation time depends on the specific reaction reagent. The staining method of the present application is applied to immunohistochemical staining, conventional staining, and special staining. In immunohistochemical staining, the primary antibody incubation, the secondary antibody incubation, the DAB staining, and the like can be applied to the method for the operation of the mixed reaction reagent disclosed in the technical scheme; likewise, the technical solution of the present application can be applied to the staining of eosin. Conventional dyeing such as hematoxylin staining Medium.
[0039] 以上所述仅为本申请的优选实施例， 并非因此限制本申请的专利范围， 凡是在 本申请的申请构思下， 利用本申请说明内容所作的技术延伸或再创造均包括在 本申请的专利保护范围内。 The above description is only a preferred embodiment of the present application, and is not intended to limit the scope of the patents of the present application, and the technical extension or re-creation made by the content of the present application is included in the present application. Within the scope of patent protection.
发明概述 Summary of invention
技术问题 technical problem
问题的解决方案 Problem solution
发明的有益效果
Advantageous effects of the invention
Claims
[权利要求 1] 一种病理切片的染色方法， 应用于自动化染色系统， 其特征在于， 包 括步骤： [Claim 1] A method for staining a pathological section, which is applied to an automated staining system, characterized in that it comprises the steps of:
b、 向病理切片加入反应试剂； c、 静置孵育 1-10分钟； b, adding a reagent to the pathological section; c, incubated for 1-10 minutes;
d、 完成步骤 c后， 对反应试剂进行混匀。 d. After completing step c, mix the reagents.
[权利要求 2] 根据权利要求 1所述病理切片的染色方法， 其特征在于， 在步骤 d之 后还包括步骤： [Claim 2] The method for staining a pathological section according to claim 1, further comprising the step of: after step d:
e、 完成步骤 d之后，至少重复一次步骤 c、 步骤 d操作。 e. After completing step d, repeat steps c and d at least once.
[权利要求 3] 根据权利要求 2所述病理切片的染色方法， 其特征在于， 染色过程中 使用载玻片盖体； 所述载玻片盖体包括反应腔、 加样槽和突起部； 载 玻片盖体放置于自动化染色仪托盘内， 盖体的反应腔位置对应载玻片 上的组织切片位置。 [Claim 3] The method for staining a pathological slice according to claim 2, wherein a slide cover is used in the dyeing process; the slide cover includes a reaction chamber, a sample introduction groove, and a protrusion; The slide cover is placed in an automated stainer tray, and the position of the reaction chamber of the cover corresponds to the position of the tissue slice on the slide.
[权利要求 4] 根据权利要求 3所述病理切片的染色方法， 其特征在于， 所述步骤 b 具体包括： 在盖体加样槽加入反应试剂。 [Claim 4] The method for staining a pathological section according to claim 3, wherein the step b specifically comprises: adding a reaction reagent to the lid sample loading tank.
[权利要求 5] 根据权利要求 4所述病理切片的染色方法， 其特征在于， 所述步骤 d 具体包括： 将盖体突出部抬起再放回原来位置， 静置载玻片后， 重复 操作， 使得反应试剂充分混匀。 [Claim 5] The method for dyeing a pathological slice according to claim 4, wherein the step d specifically comprises: lifting the lid protrusion and returning it to the original position, and repeating the operation after standing the slide , allowing the reaction reagent to mix well.
[权利要求 6] 根据权利要求 4所述病理切片的染色方法， 其特征在于， 所述步骤 d 具体包括： 将盖体突出部抬起并向突出部方向拖动后再将放置回原来 位置， 静置载玻片后， 重复操作， 使得反应试剂充分混匀。 [Claim 6] The method for dyeing a pathological slice according to claim 4, wherein the step d specifically comprises: lifting the lid protrusion and dragging it toward the protrusion, and then returning to the original position, After standing on the slide, repeat the operation to allow the reaction reagent to mix well.
[权利要求 7] 根据权利要求 2所述病理切片的染色方法， 其特征在于， 染色过程中 使用液态油膜； 所述油膜包裹于病理切片外； 油膜成分为含碳数为 C10-C16的碳氢化合物。 [Claim 7] The method for staining a pathological section according to claim 2, wherein a liquid oil film is used in the dyeing process; the oil film is wrapped outside the pathological section; and the oil film component is a hydrocarbon having a carbon number of C10-C16 Compound.
[权利要求 8] 根据权利要求 7所述病理切片的染色方法， 其特征在于， 所述步骤 b 具体包括： 向病理切片加入试剂， 试剂穿过油膜与病理切片接触， 油 膜包裹于反应试剂外。 [Claim 8] The method for staining a pathological section according to claim 7, wherein the step b specifically comprises: adding a reagent to the pathological section, the reagent is in contact with the pathological section through the oil film, and the oil film is wrapped outside the reaction reagent.
[权利要求 9] 根据权利要求 7所述病理切片的染色方法， 其特征在于， 所述步骤 d 具体包括： 向由油膜包裹的反应试剂提供气流， 气流的作用下反应试
剂在油膜范围内流动； 静置后， 重复操作， 使得反应试剂充分混匀。 [Claim 9] The method for staining a pathological slice according to claim 7, wherein the step d specifically comprises: providing a gas flow to the reaction reagent wrapped by the oil film, and reacting under the action of the gas flow The agent flows in the oil film range; after standing, the operation is repeated to allow the reaction reagent to be thoroughly mixed.
[权利要求 10] 根据权利要求 7所述病理切片的染色方法， 其特征在于， 所述油膜成 分为 C 15H 32。
[Claim 10] The method of dyeing a pathological section according to claim 7, wherein the oil film component is C 15 H 32 .
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| CN102266386A (en) * | 2010-06-01 | 2011-12-07 | 四川大千药业有限公司 | Scutellaria baicalensis extract and preparation method and uses thereof |
| CN202204717U (en) * | 2011-08-18 | 2012-04-25 | 浙江世纪康大医疗科技有限公司 | Pathological section covering plate of staining instrument |
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| JP6237194B2 (en) * | 2013-12-18 | 2017-11-29 | コニカミノルタ株式会社 | Dyeing method |
| CN108387428A (en) * | 2018-02-26 | 2018-08-10 | 深圳安诺康生物医疗有限公司 | A kind of colouring method of pathological section |
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| CN202204717U (en) * | 2011-08-18 | 2012-04-25 | 浙江世纪康大医疗科技有限公司 | Pathological section covering plate of staining instrument |
| JP6237194B2 (en) * | 2013-12-18 | 2017-11-29 | コニカミノルタ株式会社 | Dyeing method |
| CN103698507A (en) * | 2014-01-09 | 2014-04-02 | 南京医科大学第一附属医院 | Detachable and sealed space glass slide incubator |
| CN108387428A (en) * | 2018-02-26 | 2018-08-10 | 深圳安诺康生物医疗有限公司 | A kind of colouring method of pathological section |
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