Ultralow-temperature freezing and sealing type preservation method

An ultra-low temperature freezing and ultra-low temperature preservation technology, applied in the preservation, application, animal husbandry and other directions of human or animal body, can solve the problems of the difficulty of maintaining the sterile state of liquid nitrogen, cross-infection of embryos, etc., to avoid misplacement, increase The effect of space utilization and avoiding re-contamination

Active Publication Date: 2019-12-03
江苏瑞辅达医疗器械有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the liquid nitrogen produced is sterile, it is extremely difficult to maintain the sterile state of liquid nitrogen during transportation, storage and use
Moreover, the embryos of patients infected with bacteria or viruses may also indirectly transmit pathogens to other "healthy" embryos in the liquid nitrogen tank, resulting in cross-infection of embryos

Method used

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  • Ultralow-temperature freezing and sealing type preservation method
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  • Ultralow-temperature freezing and sealing type preservation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Embodiment 1: prepare aseptic refrigerant

[0043] Step 1. Provide a container B. The container B is an open structure, wherein the opening of the container B is the second opening 11 . The size of the container B can also be selected according to requirements, which is not limited in the present invention. In this embodiment, the container B is a foam box with a length of 260mm*width of 150mm*depth of 115mm without a cover. An initial cold source 12 is contained in the container B. In this embodiment, the initial cold source 12 contained in the container B is commercially available liquid nitrogen. The initial cooling source 12 has a liquid layer 12A and a vapor layer 12B above the liquid layer 12A, wherein the vapor layer 12B is formed by volatilization of the liquid layer 12A. In order to clearly show the difference between the two, the liquid layer 12A and the vapor layer 12B are drawn with different hatching in the drawings. Specifically, the depth of the cont...

Embodiment 2

[0051] Embodiment 2: Aseptic refrigerant composition and temperature measurement

[0052] The liquid air prepared in the plastic tube in Example 1 was placed at room temperature, the vaporized air was introduced into a vacuumized aluminum foil bag, and the proportion of nitrogen in the liquid air was measured by gas chromatography. Gas chromatograph adopts Agilent6890, equipped with TCD detector, carbon molecular sieve packed column, 2m×2mmID, carrier gas is nitrogen, carrier gas flow rate: 20mL / min, column temperature 40°C, run time 5min. Injection method: gas valve injection, quantitative tube volume 1mL, sample filling time 0.5min, sampling time 0.5min.

[0053] As a result of the measurement, the nitrogen content in the obtained liquid air is about 85.5%. The ultra-low temperature thermometer measures the temperature of liquid air at about -195°C. Using liquid air as a refrigerant, you can use liquid nitrogen as a cold source anytime and anywhere to prepare sterile refri...

Embodiment 3

[0054] Embodiment 3: Aseptic refrigerant contamination test

[0055] Prepare aseptic liquid air as described in Example 1. After repeatedly dipping the liquid air with a sterile cotton swab, apply the sample (experimental group) dipped by the cotton swab on a Columbia blood plate, place at 35°C, 5% CO 2 Cultivate in the incubator for 48 hours, and observe the bacterial growth. Take 10 milliliters of the initial cold source liquid nitrogen used to prepare the sterile liquid air as a control (control group). The results showed that there were two kinds of yellow and off-white colonies growing on the blood plate of the control group ( image 3 A), while the experimental group had no colony growth (attached image 3 B). Further, the two colonies of the experimental group were isolated and purified, Gram staining and molecular identification showed that the yellow colonies were Gram-positive cocci under the bacterial microscope, and were identified as Neomicrococcus aestuarii st...

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Abstract

The invention provides an ultralow-temperature freezing and sealing type preservation method which comprises the following steps: 1) providing a sterile container with an opening; 2) putting the sterile container into an initial cold source, wherein the height of the opening of the sterile container is greater than a liquid level layer of the initial cold source; 3) forming a gasification gas as asterile freezing medium in the sterile container; 4) putting a sample to be frozen on a carrier into the sterile freezing medium, and performing vitrification freezing; and 5) sealing the sterile container, and performing ultralow-temperature preservation on the sealed sterile container together with the vitrification freezing sample sealed in the sealed sterile container.

Description

technical field [0001] The invention relates to the field of biological sample preservation, in particular to an ultra-low temperature freezing and closed preservation method. Background technique [0002] Medicine, life sciences (including assisted reproduction), food, chemical industry and other fields all need cryopreservation. Cryogenic storage requires the use of cold sources, including but not limited to conventional refrigerants such as liquid nitrogen. [0003] Taking the field of human assisted reproduction as an example, this field requires cryopreservation of cell samples such as embryos and eggs, and liquid nitrogen is the main cold source. Vitrification has become the main method for freezing human embryos and eggs because of its fast cooling rate (above 20,000°C / min) and high recovery survival rate (≥95%). However, the potential contamination risk of vitrification technology has always troubled practitioners of assisted reproduction. During the vitrification...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
CPCA01N1/02A01N1/0268
Inventor 彭秋平薛松果王荣祥彭文林胥尧匡延平
Owner 江苏瑞辅达医疗器械有限公司
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