WO2023036225A1 - 用于生物组织冷冻保存或解冻复苏的装置 - Google Patents

用于生物组织冷冻保存或解冻复苏的装置 Download PDF

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WO2023036225A1
WO2023036225A1 PCT/CN2022/117749 CN2022117749W WO2023036225A1 WO 2023036225 A1 WO2023036225 A1 WO 2023036225A1 CN 2022117749 W CN2022117749 W CN 2022117749W WO 2023036225 A1 WO2023036225 A1 WO 2023036225A1
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solution
grid
carrier
tubular body
thawing
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PCT/CN2022/117749
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French (fr)
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陈昊楠
舒怡玮
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深圳拜尔洛克生物技术有限公司
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Publication of WO2023036225A1 publication Critical patent/WO2023036225A1/zh

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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/02—Preservation of living parts
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M3/00—Tissue, human, animal or plant cell, or virus culture apparatus

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  • the invention relates to the technical field of bioengineering, in particular to a device for cryopreservation or thawing recovery of biological tissues.
  • Cell cryopreservation technology is an important technology in many biological fields. It is also important to restore cells from a cryogenically frozen state to a normal metabolic state.
  • the process of thawing and resuscitation includes the process of removing the cryoprotectant and replacing it with a normal medium.
  • the devices in the prior art are complicated to operate when the embryo is frozen or thawed, and the efficiency is low.
  • the main purpose of the present invention is to provide a device for cryopreservation or thawing and resuscitating of biological tissues, so as to solve the problems of complex operation and low efficiency in the prior art when freezing or thawing embryos.
  • the device for cryopreservation or thawing and resuscitating biological tissue includes: a carrier, the carrier includes a tubular body, and one end of the tubular body is provided with a grid-like structure, and the grid-like structure prevents the biological tissue from Exit the tubular body through the solution outlet.
  • the tubular body has a solution inlet and a solution outlet, and the solution outlet is provided with the grid structure.
  • the tubular body includes a solution channel through which a solution flows, and the solution enters the solution channel through the solution inlet and exits the solution channel through the solution outlet.
  • the solution inlet is arranged at the end of the tubular body.
  • the solution outlet is arranged near the proximal end of the tubular body.
  • the grid structure includes a plurality of grid bars, and the distance between every two grid bars in the plurality of grid bars is smaller than the diameter of the biological tissue and larger than the diameter of the solution molecules.
  • the length of the grid structure is 0.8-1.5 mm.
  • the embryos are restricted by the grid structure at the proximal end of the carrier and will not float out, so that the delivery or removal of cryoprotectants can be carried out in an orderly manner, effectively improving It improves the efficiency of embryo/egg cryopreservation or thawing recovery operation.
  • FIG. 1 is a schematic perspective view of a carrier according to an embodiment of the present invention
  • Figure 2 is a side view of a carrier according to an embodiment of the present invention.
  • Fig. 3 is a schematic cross-sectional view of line A-A in Fig. 2 .
  • a device for cryopreservation or thawing and resuscitation of biological tissue is provided.
  • the inner diameter can be 0.8-1.5 mm.
  • the solution passage for the solution to flow in the tubular main body 11, the solution passage includes a solution inlet 12 arranged at the end of the tubular main body 11 and a solution outlet 13 arranged near the proximal end of the tubular main body 11, and the solution can enter the solution through the solution inlet 12 channel and exit the solution channel through the solution outlet 13.
  • the solution channel of the carrier can carry biological tissue, and one end of the tubular body 11 of the carrier is provided with a grid-like structure 15, specifically, a grid-like structure 15 is arranged at the solution outlet 13, and the grid-like structure 15 includes a plurality of grid bars.
  • the slit spacing between every two grid bars in each grid bar is smaller than the diameter of the biological tissue and larger than the diameter of the solution molecule, wherein the slit spacing (grid pitch) can be 0.05-0.08 mm.
  • the biological tissue may be biological materials such as embryos, eggs, cells, etc., which is not limited in the present application, and the following description will be made by taking the biological tissue as an embryo as an example.
  • a glass capillary can be used to deliver embryos to the carrier.
  • the bars of the grid-like structure 15 are elastic, and the grid-like structure 15 can be deformed when in contact with a foreign object (such as a glass capillary), so that the foreign object can pass through the grid-like structure 15 .
  • a foreign object such as a glass capillary
  • the glass capillary is used to put the embryo into the carrier, after the glass capillary touches the grid-like structure 15, the grid-like structure 15 will be deformed, and if the glass capillary is continued to be inserted, the glass capillary can pass through the grid-like structure 15 and enter the solution channel. The embryos in the glass capillary are dropped into the solution channel.
  • the grid-like structure 15 After the embryos are put in, the grid-like structure 15 returns to its original state after the glass capillary leaves the solution channel. Since the slit spacing of the grid-like structure 15 is smaller than the diameter of the embryo, the embryo can be restricted in the solution channel (the proximal end of the carrier) , and will not leave the carrier with the flow of liquid. In addition, embryos can also be placed from the end of the carrier (solution inlet). Specifically, the solution channel of the carrier is filled with basic culture solution, and then a glass capillary is used to insert the carrier from the end of the carrier, and then the embryos are released. When a pipette pump or an air pump is used to pour the solution into the carrier, the solution will bring the embryo to the proximal position of the carrier. Since the slit spacing of the grid structure 15 is smaller than the diameter of the embryo, the embryo can be limited to the proximal position of the carrier. end, and will not leave the carrier with the flow of liquid.
  • the solution leaves the carrier through the grid-like structure, and the proximal end of the carrier can be placed in a container to collect waste liquid (usually tens to hundreds of microliters of solution).
  • waste liquid usually tens to hundreds of microliters of solution.
  • thawing application operations When performing thawing application operations, first prepare a plate of thawing solution that has been preheated to 37 degrees, then take the carrier out of the liquid nitrogen, and immerse the proximal part of the carrier into the thawing solution that has been preheated to 37 degrees, after 50 seconds , connect the hose to the end of the carrier, and then use an automated solution-propelling instrument (such as a syringe pump or an air pump) to slowly pour the diluent and cleaning solution from the solution inlet into the channel of the carrier according to an automatic procedure.
  • an automated solution-propelling instrument such as a syringe pump or an air pump
  • the cryoprotectant in the embryo and near the embryo is taken away by the thawing liquid to achieve the purpose of thawing and resuscitating, replacing the existing manual operation scheme.
  • the solution leaves the carrier through the grid-like structure, and the proximal end of the carrier can be placed in a container to collect waste liquid (usually tens to hundreds of microliters of solution).
  • waste liquid usually tens to hundreds of microliters of solution.
  • the grid structure restricts the embryos to the proximal end of the carrier and prevents them from floating out, so that the delivery or removal of the cryoprotectant can be carried out in an orderly manner, Effectively improve the efficiency of embryo cryopreservation or thawing recovery operation.

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Abstract

一种用于生物组织冷冻保存或解冻复苏的装置，其包括：载体，所述载体包括管状主体，所述管状主体的一端设置有栅状结构，所述栅状结构阻止生物组织通过所述溶液出口离开所述管状主体。所述装置能够有序进行冷冻保护剂的投送或移除，有效提高了胚胎/卵子冷冻保存或解冻复苏操作的效率。

Description

用于生物组织冷冻保存或解冻复苏的装置 技术领域
本发明涉及生物工程技术领域，尤其涉及一种用于生物组织冷冻保存或解冻复苏的装置。
背景技术
细胞冷冻保存技术在很多生物领域是一项重要的技术，把细胞从低温冷冻状态恢复到正常代谢状态也同样重要，解冻复苏的过程包含了把冷冻保护剂移除替换成正常培养基的过程。
现有技术的装置在胚胎冷冻或解冻时操作复杂，效率较低。
发明内容
本发明的主要目的在于提供一种用于生物组织冷冻保存或解冻复苏的装置，以解决现有技术的装置在进行胚胎冷冻或解冻时存在的操作复杂、效率较低等问题。
根据本发明实施例的用于生物组织冷冻保存或解冻复苏的装置，其包括：载体，所述载体包括管状主体，所述管状主体的一端设置有栅状结构，所述栅状结构阻止生物组织通过所述溶液出口离开所述管状主体。
其中，所述管状主体具有溶液入口和溶液出口，所述溶液出口设置有所述栅状结构。
其中，所述管状主体包括供溶液流动的溶液通道，溶液通过所述溶液入口进入所述溶液通道并通过所述溶液出口离开所述溶液通道。
其中，所述溶液入口设置在所述管状主体的末端。
其中，所述溶液出口设置在所述管状主体的近端附近。
其中，所述栅状结构包括多个栅条，所述多个栅条中的每两个栅条之间的距离小于所述生物组织的直径且大于溶液分子的直径。
其中，所述栅状结构的长度为0.8-1.5毫米。
根据本发明的技术方案，在进行冷冻保存或解冻复苏操作时，胚胎被栅状结构限制在载体的近端不会飘出，这样能够有序进行冷冻保护剂的投送或移除，有效提高了胚胎/卵子冷冻保存或解冻复苏操作的效率。
附图说明
此处所说明的附图用来提供对本发明的进一步理解，构成本申请的一部分，本发明的示意性实施例及其说明用于解释本发明，并不构成对本发明的不当限定。在附图中：
图1是根据本发明实施例的载体的立体示意图；
图2是根据本发明实施例的载体的侧视图；
图3是图2中A-A线的剖面示意图。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚，下面将结合本发明具体实施例及相应的附图对本发明技术方案进行清楚、完整地描述。显然，所描述的实施例仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。
以下结合附图，详细说明本发明各实施例提供的技术方案。
根据本发明实施例提供一种用于生物组织冷冻保存或解冻复苏的装置，参考图1至图3，所述装置包括载体，所述载体具有呈管状的主体11，管状主体11的外径可为1.5-2.5毫米、内径可为0.8-1.5毫米。管状主体11内具有供溶液流动的溶液通道，该溶液通道包括设置在管状主体11的末端的溶液入口12和设置在管状主体11的近端附近的溶液出口13，溶液可以通过溶液入口12进入溶液通道并通过溶液出口13离开溶液通道。
载体的溶液通道可承载生物组织，载体的管状主体11的一端设置有栅状结构15，具体来说，在溶液出口13设置有栅状结构15，栅状结构15包括 多个栅条，这些多个栅条中的每两个栅条之间的细缝间距小于生物组织的直径且大于溶液分子的直径，其中所述细缝间距(栅距)可为0.05-0.08毫米。这样，当溶液(冷冻液或解冻液)在溶液通道内由末端向近端流动的情况下，溶液推动生物组织到载体的近端位置，溶液通过栅状结构15离开载体，而生物组织则被栅状结构15阻挡在载体的近端的位置附近，防止生物组织随着溶液离开载体。其中，所述生物组织可以是胚胎、卵子、细胞等生物材料，本申请对此不进行限制，下面以生物组织是胚胎为例进行描述。
在本申请实施例中，可使用玻璃毛细管向载体投放胚胎。栅状结构15的栅条具有弹性，栅状结构15可在与外物(例如玻璃毛细管)接触时发生形变从而使得外物穿过栅状结构15。使用玻璃毛细管向载体投放胚胎时，玻璃毛细管触碰到栅状结构15后，栅状结构15会发生形变，继续插入玻璃毛细管，则玻璃毛细管就能够穿过栅状结构15进入溶液通道中，将玻璃毛细管内的胚胎投放到溶液通道中。待胚胎投放后，玻璃毛细管离开溶液通道后栅状结构15则恢复到原来的状态，由于栅状结构15的细缝间距小于胚胎的直径，胚胎可以被限制在溶液通道中(载体的近端)，并不会随着液体的流动离开载体。此外，还可从载体的末端(溶液入口)投放胚胎，具体地，先将载体的溶液通道内填满基础培养液，然后使用玻璃毛细管从载体的末端插入载体，再放出胚胎。当使用移液泵或者气压泵往载体灌入溶液时，溶液会把胚胎带到载体的近端位置，由于栅状结构15的细缝间距小于胚胎的直径，胚胎可以被限位在载体的近端，并不会随着液体的流动离开载体。
当进行冷冻应用操作时，首先使用移液枪把载体的溶液通道内部灌满基础培养液，然后把载体平放在显微镜上，使用玻璃毛细管把胚胎从培养皿取出，穿过载体近端的栅状结构，将胚胎投放到载体中，然后把载体的末端接上软管，使用自动化的溶液推动仪器(例如注射泵或者气压泵)，把冷冻液慢慢从溶液入口灌入载体的溶液通道。溶液经过溶液通道流动到载体近端时，溶液内的冷冻保护剂就可以扩散到胚胎内部，达到冷冻保护剂投送的目的，替代现有的人工操作的方案。溶液经过栅状结构离开载体，载体的近端可以放在一个容器内，收集废液(一般为几十到几百微升的溶液)。当处理完成后，使用者把载体从软管拔出，投入液氮保存。
当进行解冻应用操作时，先准备一盘已经预热到37度的解冻液，然后把载体从液氮取出，把载体近端的部分侵入已经预热到37度的解冻液里，50秒后，把软管接上载体的末端，然后使用自动化的溶液推动仪器(例如注射泵或者气压泵)，把稀释液和清洗液按自动程序慢慢从溶液入口灌入载体的通道。溶液经过载体的溶液通道流动到载体近端时，胚胎体内和胚胎附近的冷冻保护剂被解冻液带走移除，达到解冻复苏的目的，替代现有的人工操作的方案。溶液经过栅状结构离开载体，载体的近端可以放在一个容器内，收集废液(一般为几十到几百微升的溶液)。当处理完成后，使用者把载体平放于显微镜上，将玻璃毛细管穿过栅状结构将胚胎取出，放置到培养皿内继续培养。
通过本申请的上述实施例，在进行冷冻保存或解冻复苏操作时，栅状结构将胚胎限制在载体的近端而不能够飘出，从而能够有序进行冷冻保护剂的投送或移除，有效提高了胚胎冷冻保存或解冻复苏操作的效率。
以上所述仅为本发明的实施例而已，并不用于限制本发明，对于本领域的技术人员来说，本发明可以有各种更改和变化。凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的权利要求范围之内。

Claims (7)

  1. 一种用于生物组织冷冻保存或解冻复苏的装置，其特征在于，包括：
    载体，所述载体包括管状主体，所述管状主体的一端设置有栅状结构，所述栅状结构用于阻挡生物组织离开所述管状主体。
  2. 根据权利要求1所述的装置，其特征在于，所述管状主体具有溶液入口和溶液出口，所述溶液出口设置有所述栅状结构。
  3. 根据权利要求2所述的装置，其特征在于，所述管状主体包括供溶液流动的溶液通道，溶液通过所述溶液入口进入所述溶液通道并通过所述溶液出口离开所述溶液通道。
  4. 根据权利要求2所述的装置，其特征在于，所述溶液入口设置在所述管状主体的末端。
  5. 根据权利要求2所述的装置，其特征在于，所述溶液出口设置在所述管状主体的近端附近。
  6. 根据权利要求1所述的装置，其特征在于，所述栅状结构包括多个栅条，所述多个栅条中的每两个栅条之间的距离小于所述生物组织的直径且大于溶液分子的直径。
  7. 根据权利要求1至6中任一项所述的装置，其特征在于，所述栅状结构的长度为0.8-1.5毫米。
PCT/CN2022/117749 2021-09-10 2022-09-08 用于生物组织冷冻保存或解冻复苏的装置 WO2023036225A1 (zh)

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