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Pancreatic island freezing protective agent and using method thereof

A technology of cryoprotectant and islet, which is applied in the field of islet cryoprotectant, can solve the problem of ineffective protection of islet cell clusters, and achieve the effect of inhibiting degradation and inhibiting the action of digestive enzymes

Inactive Publication Date: 2011-09-28
FUZHOU GENERAL HOSPITAL OF NANJING MILITARY COMMAND P L A
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The cryoprotectant and cryopreservation method of the conventional single cell suspension can not effectively protect the islet cell mass

Method used

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  • Pancreatic island freezing protective agent and using method thereof
  • Pancreatic island freezing protective agent and using method thereof
  • Pancreatic island freezing protective agent and using method thereof

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0031] Reagent source:

[0032] M-199 medium is a commercial product purchased from HYCLONE; AB serum is from healthy human serum in a blood bank; 1M Hepes is a commercial product purchased from Mediatech Cellgro, USA; Ulinastatin is a clinically used drug for injection; DMSO was a commercial product of clinical use, which was purchased from Sigma, USA; isopropanol was an analytical grade chemical commodity, which was purchased from Shanghai Chemical Reagent Factory.

[0033] Reagent preparation:

[0034] The islet cryopreservation agent of the present invention comprises A liquid, B liquid, C liquid and D liquid, and composition is as follows:

[0035] Liquid A: basic freezing medium (500ml):

[0036] final concentration volume

[0037] M-199 Medium - 337.5ml

[0038] AB Serum 30% 150ml

[0039] Hepes (1M) 25mM 12.5ml

[0040] Ulinastatin 200IU / ml 100,000IU

[0041] Solution B: 2M DMSO medium (100ml):

[0042] final concentration volume

[0043] DMSO 2M 14.2ml

[00...

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Abstract

The invention provides a pancreatic island freezing protective agent and a using method thereof. The pancreatic island freezing protective agent comprises liquid A, liquid B, liquid C and liquid D, wherein the liquid A is a basic freezing culture medium and consists of 50 to 70 volume percent of M-199 culture medium, 30 to 50 volume percent of AB serum, 1 to 3 volume percent of 1MHepes and 100 to200 IU / ml of Ulinastatin; the liquid B is a 2MDMSO freezing culture medium and comprises 14.2 volume percent of DMSO and 85.8 volume percent of basic freezing culture medium; the liquid C is a 3MDMSOfreezing culture medium and comprises 1.3 volume percent of DMSO2 and 78.7 volume percent of basic freezing culture medium; and the liquid D is isopropanol. The pancreatic island is resuspended by a basic culture medium of high-concentration serum, and different concentrations of DMSO freezing liquid is added gradually incrementally, so that the DMSO can be permeated into pancreatic island cell masses. The isopropanol serves as a freezing medium and has the effect of reducing temperature slowly. The pancreatic island freezing protective agent is used for effectively preserving islet cells obtained through separation, so that the islet cells are prevented from being damaged by freezing and are provided for clinical application.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a protective agent for cryopreservation of pancreatic islets and its application method. Background technique [0002] Diabetes seriously affects people's health. Diabetes is divided into Type I and Type II. Type 1 diabetes is insulin-dependent, and approximately 10% of diabetics have type 1 diabetes. About 30% of type II diabetes patients will also develop into insulin-dependent type in the middle and late stages (duration of more than 20 years). Insulin injection is currently an effective means of controlling blood sugar in diabetic patients, but exogenous insulin cannot achieve the purpose of physiologically regulating blood sugar, so it cannot prevent the occurrence and development of diabetic complications such as nephropathy, nervous system disease, retinopathy and cardiovascular disease , thereby greatly reducing the patient's quality of life and eventual...

Claims

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Application Information

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IPC IPC(8): A01N1/02
Inventor 陈津黄梁浒马予洁王庆华谭建明
Owner FUZHOU GENERAL HOSPITAL OF NANJING MILITARY COMMAND P L A
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