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A skin cryoprotectant and skin preservation method

A cryogenic and protective agent technology, applied in the field of cryogenic preservation of biological materials, can solve the problems of patient infection risk, difficult quality assurance, component differences, etc., to avoid infection risks, improve success rate, and achieve the effect of small changes in biological properties

Active Publication Date: 2021-08-31
SHANDONG UNIV QILU HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The main defect of cryopreservation of skin at present is that fetal bovine serum is usually added as a cryoprotectant component, but because it is a product of animal origin, the patient has a certain risk of infection when the skin is replanted in cryopreservation
However, due to differences in bee species, bee sources, and environments, the components of honeybees vary greatly, and the quality is difficult to guarantee, and it is difficult to grasp in practice.

Method used

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  • A skin cryoprotectant and skin preservation method
  • A skin cryoprotectant and skin preservation method
  • A skin cryoprotectant and skin preservation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1 preparation cryoprotectant and skin preservation

[0030] Preparation of cryoprotectant: Mix 80ml of RPMI 1640 medium (Hyclone, USA) with 10ml of human serum albumin (BR, China), then slowly add 10ml of dimethyl sulfoxide (Sigma, USA) to prepare the base solution , and then add maltodextrin and urea (10 kinds of cryoprotectants were prepared because of the different final concentrations of maltodextrin and urea, and the final concentrations of maltodextrin and urea in each cryoprotectant are shown in Table 1) , fully dissolved and mixed to prepare a cryoprotectant, and placed in a 4°C refrigerator for pre-cooling.

[0031] Prepare Kreb's Ringer's phosphate buffer: get 11.421g disodium hydrogen phosphate (Sigma, U.S.), 2.26g sodium dihydrogen phosphate (Sigma, U.S.), and dilute to 500ml with distilled water to prepare 0.1M phosphate buffer; 0.9g sodium chloride (Sigma, the U.S.), 1.15g potassium chloride (Sigma, the U.S.), 1.22g calcium chloride (Sigma, the...

experiment example 1

[0040] Experimental Example 1 Histomorphological Observation

[0041] Paraffin sections were made by HE staining method for histomorphological observation.

[0042] After rapid rewarming in a water bath at 42°C, put the pigskin preserved in deep cryogenic temperature (0.75% urea + 15% maltodextrin group) and the pigskin preserved in glycerol in 4% formaldehyde, and fix them overnight in a refrigerator at 4°C. The samples were sent to the Department of Pathology, East District, Shandong Provincial Hospital of Traditional Chinese Medicine (No. 16369, Jingshi Road, Lixia District, Jinan City, Shandong Province) for HE-stained paraffin sections.

[0043] The paraffin sections of the samples were observed and photographed with an optical microscope (Olympus BX53), and the results were as follows: figure 1 and figure 2 shown. Depend on figure 1 and figure 2 It can be seen from the comparison of the morphological results that, compared with the control example, the pigskin tis...

experiment example 2

[0044] Experimental Example 2 Survival rate of pig skin slices after resuscitation

[0045] Use the succinate dehydrogenase assay to detect the survival rate of pig skin slices after resuscitation, the steps are as follows:

[0046] (1) Trim the rewarmed pigskin to a size of about 1cm 2 Two skin slices were taken for each treatment period in each group, and the quality of the skin slices was measured.

[0047] (2) Put 2 pieces of skin into a 5ml syringe (Weigao, China), apply Vaseline on the back of the syringe to seal it, then inhale 1ml of the mixed reaction solution into the syringe, try to remove air bubbles, and place the syringe needle with Rubber stopper seal.

[0048] (3) Place the sealed syringe in a constant temperature incubator (Thermo Fisher, USA) at 37°C and incubate for 60 min, and a red precipitate can be observed on the skin slice.

[0049] (4) After removing the reaction solution, inhale 2ml of ethylene glycol ether into each syringe, and place it at room ...

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Abstract

The invention discloses a deep cryoprotectant for skin, which mainly consists of dimethyl sulfoxide, cell culture medium, human serum albumin, maltodextrin and urea, wherein dimethyl sulfoxide, cell culture medium, human blood Albumin constitutes the base fluid, in which dimethyl sulfoxide accounts for 5% to 12%, human serum albumin accounts for 5% to 10%, and the balance is cell culture medium, calculated by volume percentage; maltodextrin The concentration is 10%-20%, and the concentration of urea is 0.5%-1.5%. The skin cryoprotectant of the invention can be used for cryoprotection of human allogeneic skin. The cell survival rate of human allogeneic skin preserved by the preservation method of the present invention is high, and the biological properties of the skin graft have little change, which is conducive to the rapid generation of granulation tissue at the transplantation site of the patient, improves the success rate of replantation, and shortens the treatment time of the patient at the same time. time and reduce treatment costs.

Description

technical field [0001] The invention relates to a deep cryoprotectant for skin and a skin preservation method using maltodextrin and urea as raw materials, and belongs to the technical field of low temperature preservation of biological materials. Background technique [0002] The skin is the largest organ that covers the entire human body surface to prevent the loss of body fluids, prevent harmful substances and microorganisms from invading from the outside, resist physical and chemical stimuli, etc., and perform the function of protecting the body. In recent years, the number of people injured due to accidents such as car accidents and burns has increased year by year, and skin detachment of limbs and body parts has occurred from time to time. [0003] Currently, there are three main methods for treating damaged skin tissue: autografting of the patient's own skin, allografting of other people's skin, and xenografting of animal-derived skin. Allografts, which primarily aid...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221A01N1/0226
Inventor 朱磊李夏臧传宝林俊豪王俊涛
Owner SHANDONG UNIV QILU HOSPITAL
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