CN107114356A - A kind of placenta and umbilical cord cells protect liquid - Google Patents
A kind of placenta and umbilical cord cells protect liquid Download PDFInfo
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- CN107114356A CN107114356A CN201610625635.XA CN201610625635A CN107114356A CN 107114356 A CN107114356 A CN 107114356A CN 201610625635 A CN201610625635 A CN 201610625635A CN 107114356 A CN107114356 A CN 107114356A
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- mem
- parts
- umbilical cord
- aqueous solution
- placenta
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Classifications
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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
- A01N1/10—Preservation of living parts
- A01N1/12—Chemical aspects of preservation
- A01N1/122—Preservation or perfusion media
- A01N1/126—Physiologically active agents, e.g. antioxidants or nutrients
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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
- A01N1/10—Preservation of living parts
- A01N1/12—Chemical aspects of preservation
- A01N1/122—Preservation or perfusion media
- A01N1/124—Disinfecting agents, e.g. antimicrobials
-
- 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
- A01N1/10—Preservation of living parts
- A01N1/12—Chemical aspects of preservation
- A01N1/122—Preservation or perfusion media
- A01N1/125—Freeze protecting agents, e.g. cryoprotectants or osmolarity regulators
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Dentistry (AREA)
- General Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
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- Cosmetics (AREA)
Abstract
本发明提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂和氨基糖苷类抗生素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10‑30mg、所述氨基糖苷类抗生素180‑240U，所述MEM–α培养基水溶液的浓度为10‑15mg/mL。本发明提供的保护液不仅能够有效杀死或抑制支原体的形成，防止胎盘细胞或脐带细胞感染，而且通过加入保护成分能够为胎盘或脐带细胞提供一个稳定的渗透压环境，有效模拟人体环境，提高细胞生存率，同时能够为细胞短时间内保存和运输提供营养成分，提高细胞的免疫性能，防止细胞感染，有效提高细胞的生存力，延长细胞运输时间。The invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors and aminoglycoside antibiotics with MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is respectively Dissolve 10-30 mg of the mycoplasma inhibitor, 180-240 U of the aminoglycoside antibiotics, and the concentration of the MEM-α medium aqueous solution is 10-15 mg/mL. The protective solution provided by the invention can not only effectively kill or inhibit the formation of mycoplasma, prevent infection of placental cells or umbilical cord cells, but also provide a stable osmotic pressure environment for placental or umbilical cord cells by adding protective components, effectively simulating the human body environment, and improving Cell survival rate, at the same time, it can provide nutrients for cell storage and transportation in a short time, improve the immune performance of cells, prevent cell infection, effectively improve cell viability, and prolong cell transportation time.
Description
技术领域technical field
本发明属于胎盘及脐带细胞保存技术领域，特别涉及一种胎盘及脐带细胞保护液。The invention belongs to the technical field of placenta and umbilical cord cell preservation, in particular to a placenta and umbilical cord cell protection solution.
背景技术Background technique
近年来，保存胎盘干细胞和脐带血干细胞的热潮逐年上涨，大多数人已经知晓利用脐带血干细胞能够治愈血液系统疾病等等诸多好处，但是对于胎盘干细胞的生物医学领域的重大用途却知之甚少。保存胎盘主要是从保存的胎盘中提取间充质干细胞，胎盘中含有的间充质干细胞非常丰富，这种间充质干细胞属于多能干细胞，具有强大的增殖能力和多向分化潜能，能够培养发育成人体的各种组织器官与神经系统，此外，胎盘间充质干细胞治疗疾病范围比较广泛，例如治疗心、脑血管疾病、神经系统疾病、肝脏疾病、骨组织病、角膜损失、烧伤烫伤、肌病等多种肌病；同理，很多人们保存脐带血主要是从保存的脐带血中提取脐带间充质干细胞，脐带间充质干细胞具有多向分化潜能、造血支持和促进干细胞植入、免疫调控和自我复制等特点，脐带间充质干细胞是用于衰老和病变引起的组织器官损伤修复理想的种子细胞。为此，由于胎盘干细胞和脐带血干细胞对于治疗人体疾病的重要性，越来越多的人开始储存脐带间充质干细胞和胎盘干细胞以备将来不时之需。In recent years, the upsurge of preserving placental stem cells and umbilical cord blood stem cells has been increasing year by year. Most people already know that using umbilical cord blood stem cells can cure blood system diseases and many other benefits, but little is known about the important uses of placental stem cells in the biomedical field. Preserving the placenta is mainly to extract mesenchymal stem cells from the preserved placenta. The mesenchymal stem cells contained in the placenta are very rich. In addition, placental mesenchymal stem cells can treat a wide range of diseases, such as heart disease, cerebrovascular disease, nervous system disease, liver disease, bone tissue disease, corneal loss, burns, Myopathy and other myopathy; similarly, many people preserve umbilical cord blood mainly to extract umbilical cord mesenchymal stem cells from the preserved umbilical cord blood. Umbilical cord mesenchymal stem cells have multidirectional differentiation potential, hematopoietic support and promotion of stem cell implantation, With the characteristics of immune regulation and self-replication, umbilical cord mesenchymal stem cells are ideal seed cells for the repair of tissue and organ damage caused by aging and disease. For this reason, due to the importance of placental stem cells and umbilical cord blood stem cells in the treatment of human diseases, more and more people have begun to store umbilical cord mesenchymal stem cells and placental stem cells for future contingencies.
储存脐带间充质干细胞和胎盘干细胞一般包括采集、运输、交接、检测、分离、冻存、复苏、原代培养和传代培养等诸多步骤，在脐带和胎盘采集后到分离前需要运输、交接和检测三个步骤，实际操作中需要存放较长的时间。脐带和胎盘一旦采集，脱离了原有的体内环境，其细胞的活性在短时间内会迅速下降，这直接影响了其分离得到的胎盘细胞和脐带细胞的质量和活力，为此，如何维持离体胎盘和脐带细胞的活性成为了首要解决的问题。诸如时间、温度、渗透压等。The storage of umbilical cord mesenchymal stem cells and placental stem cells generally includes many steps such as collection, transportation, handover, detection, separation, cryopreservation, recovery, primary culture and subculture, etc. After the umbilical cord and placenta are collected and before they are separated, transportation, handover and There are three steps to detect, and it needs to be stored for a long time in actual operation. Once the umbilical cord and placenta are collected and separated from the original in vivo environment, the activity of the cells will drop rapidly in a short period of time, which directly affects the quality and vitality of the isolated placental cells and umbilical cord cells. Therefore, how to maintain the isolated The activity of somatic placenta and umbilical cord cells became the primary problem to be solved. Such as time, temperature, osmotic pressure, etc.
为了实现能够将胎盘和脐带进行有效保存，现有专利公开号为CN105028388A公开了保护液及其制备方法，该保护液中含有右旋糖酐、氯化钠、氯化钙、氯化钾、葡萄糖和人血白蛋白组成的溶液，为细胞提供了适宜保存环境，但是该保护液配制复杂，而且溶液稳定性差，并不能实质模拟人体环境，而且细胞冻存过程中渗透压及冻存温度均能够造成细胞存储期间活性下降，活性物质损失较快，细胞在长期存储过程中很容易凋亡，为此，急需开发一种能够延长细胞保存期且能够有效保证细胞活性，防止细胞凋亡的胎盘及脐带细胞保护液。In order to effectively preserve the placenta and umbilical cord, the existing patent publication number is CN105028388A which discloses a protection solution and a preparation method thereof. The protection solution contains dextran, sodium chloride, calcium chloride, potassium chloride, glucose and human blood The solution composed of albumin provides a suitable storage environment for the cells, but the preparation of the protective solution is complicated, and the solution stability is poor, and it cannot substantially simulate the human body environment, and the osmotic pressure and freezing temperature during the cryopreservation process of cells can cause cell storage. During the period, the activity decreases, the loss of active substances is faster, and the cells are prone to apoptosis during long-term storage. Therefore, it is urgent to develop a placenta and umbilical cord cell protection that can prolong the storage period of cells and can effectively ensure cell activity and prevent cell apoptosis. liquid.
发明内容Contents of the invention
为了解决现有技术中的保护液虽然能够保存胎盘或脐带细胞，但是这种保护液配制复杂，而且溶液稳定性差，并不能实质模拟人体环境，而且细胞冻存过程中渗透压及冻存温度均能够造成细胞存储期间活性下降，活性物质损失较快，细胞在长期存储过程中很容易凋亡等问题，本发明提供了一种胎盘及脐带细胞保护液。In order to solve the problem that although the protective solution in the prior art can preserve placental or umbilical cord cells, the preparation of this protective solution is complicated, and the solution stability is poor, and it cannot substantially simulate the human body environment. It can cause problems such as decreased activity of cells during storage, rapid loss of active substances, and easy apoptosis of cells during long-term storage. The invention provides a protective solution for placenta and umbilical cord cells.
本发明具体技术方案如下：Concrete technical scheme of the present invention is as follows:
本发明提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂和氨基糖苷类抗生素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10-30mg、所述氨基糖苷类抗生素180-240U，所述MEM–α培养基水溶液的浓度为10-15mg/mL。The invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors and aminoglycoside antibiotics with MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is respectively 10-30 mg of the mycoplasma inhibitor and 180-240 U of the aminoglycoside antibiotics are dissolved, and the concentration of the MEM-α medium aqueous solution is 10-15 mg/mL.
本发明提供的保护液通过在MEM–α培养基水溶液中加入支原体抑制剂和氨基糖苷类抗生素，为胎盘干细胞和脐带间充质干细胞提供了适宜的保存环境，通过大量实验验证，两种成分协同配伍溶解在MEM–α培养基水溶液中组成的保护液能够模拟人体环境，更加适宜胎盘干细胞和脐带间充质干细胞保存，在运输过程中起到保护细胞活性的作用，延长了运输、存储时间，防止细胞离体后在运输和存储过程中细胞凋亡，为胎盘或脐带细胞后期存储提供了有效基础。The protective solution provided by the present invention provides a suitable preservation environment for placental stem cells and umbilical cord mesenchymal stem cells by adding mycoplasma inhibitors and aminoglycoside antibiotics to the MEM-α medium aqueous solution. Through a large number of experiments, the two components synergize Compatible and dissolved in the MEM-α medium aqueous solution, the protective solution can simulate the human body environment, and is more suitable for the preservation of placental stem cells and umbilical cord mesenchymal stem cells. It can protect the cell activity during transportation and prolong the transportation and storage time. Preventing cell apoptosis during transportation and storage after isolation provides an effective basis for the later storage of placenta or umbilical cord cells.
本发明提供的每升所述MEM–α培养基水溶液包括聚乙烯吡咯烷酮100g、还原型辅酶II 5g、DMEM液体培养基500ml、三(羟甲基)甲基甘氨酸25g、地塞米松100mg、考马斯亮兰R-250 5g、3-[3-(胆酰胺基丙基)二甲氨基]丙磺酸盐5g、曲利苯兰25g、N-十二烷基肌氨酸钠100G、葡聚糖凝胶G-25[中]25g、胃蛋白酶[1:3000]25g、透明质酸酶100mg、G-418 100mg、DMEM/F12(1:1)液体培养基500ml、异丙基-β-D-硫代半乳糖苷1g、包埋剂118ml、一步法总RNA提取试剂200ml、噻孢霉素100mg、丙酮酸钠10g、精胺1g、亚精胺[精眯]1g、澳洲胎牛血清500ml、胎牛血清200ml、蛋白酶K 100mg、L-缬氨酸25g、L-亮氨酸100g、L-色氨酸10g、维生素C 25g、胸腺嘧啶核苷5g、L-谷胱甘肽5g、腺苷-5'-三磷酸二钠1g、氨苄青霉素1g、3,3'-二氨基联苯胺10g、N-(2-羟乙基)哌嗪-N'-2-乙烷磺酸25g。Every liter of the MEM-alpha medium aqueous solution provided by the present invention includes polyvinylpyrrolidone 100g, reduced coenzyme II 5g, DMEM liquid medium 500ml, tris(hydroxymethyl)methylglycine 25g, dexamethasone 100mg, coomassie Blue R-250 5g, 3-[3-(cholamidopropyl) dimethylamino] propane sulfonate 5g, Tribenilan 25g, N-Lauryl Sarcosinate 100G, Dextran Glue G-25 [medium] 25g, pepsin [1:3000] 25g, hyaluronidase 100mg, G-418 100mg, DMEM/F12 (1:1) liquid medium 500ml, isopropyl-β-D- Thiogalactoside 1g, embedding agent 118ml, one-step total RNA extraction reagent 200ml, thiosporin 100mg, sodium pyruvate 10g, spermine 1g, spermidine [spermidine] 1g, Australian fetal bovine serum 500ml, Fetal bovine serum 200ml, proteinase K 100mg, L-valine 25g, L-leucine 100g, L-tryptophan 10g, vitamin C 25g, thymidine 5g, L-glutathione 5g, adenosine -1 g of disodium 5'-triphosphate, 1 g of ampicillin, 10 g of 3,3'-diaminobenzidine, and 25 g of N-(2-hydroxyethyl)piperazine-N'-2-ethanesulfonic acid.
上述所述的DMEM(Dulbecco’s modified eagle medisum)液体培养基为一种含各种氨基酸和葡萄糖的培养基。每升该培养基内各成分及含量如下：无水氯化钙265mg/L、九水硝酸铁0.1mg/L、氯化钾400mg/L、无水硫酸镁97.6mg/L 7、氯化钠6400mg/L、无水磷酸二氢钠109mg/L、丁二酸75mg/L、丁二酸钠100mg/L、L-盐酸精氨酸84mg/L、L-盐酸胱氨酸63mg/L、甘氨酸30mg/L、L-盐酸组氨酸42mg/L、L-异亮氨酸105mg/L、L-丝氨酸42mg/L、L-苏氨酸95mg/L、L-色氨酸16mg/L、L-酪氨酸72mg/L、L-缬氨酸94mg/L、D-泛酸钙4mg/L、酒石酸胆碱7.2mg/L、叶酸4mg/L、肌醇7.2mg/L、烟酰胺4mg/L、核黄素0.4mg/L、盐酸硫胺4mg/L、盐酸吡哆辛4mg/L。进一步的改进，DMEM培养基为低糖型DMEM培养基。The above-mentioned DMEM (Dulbecco's modified eagle medisum) liquid medium is a medium containing various amino acids and glucose. The components and content of each liter of the medium are as follows: anhydrous calcium chloride 265mg/L, ferric nitrate nonahydrate 0.1mg/L, potassium chloride 400mg/L, anhydrous magnesium sulfate 97.6mg/L 7, sodium chloride 6400mg/L, anhydrous sodium dihydrogen phosphate 109mg/L, succinic acid 75mg/L, sodium succinate 100mg/L, L-arginine hydrochloride 84mg/L, L-cystine hydrochloride 63mg/L, glycine 30mg/L, L-histidine hydrochloride 42mg/L, L-isoleucine 105mg/L, L-serine 42mg/L, L-threonine 95mg/L, L-tryptophan 16mg/L, L - Tyrosine 72mg/L, L-valine 94mg/L, D-calcium pantothenate 4mg/L, choline bitartrate 7.2mg/L, folic acid 4mg/L, inositol 7.2mg/L, nicotinamide 4mg/L , riboflavin 0.4mg/L, thiamine hydrochloride 4mg/L, pyridoxine hydrochloride 4mg/L. As a further improvement, the DMEM medium is a low-sugar DMEM medium.
进一步的，所述氨基糖苷类抗生素为庆大霉素、卡那霉素、阿米卡星或妥布霉素中的一种或多种，优选的，所述氨基糖苷类抗生素由庆大霉素和卡那霉素按照5:2的重量份数比组成。Further, the aminoglycoside antibiotics are one or more of gentamicin, kanamycin, amikacin or tobramycin, preferably, the aminoglycoside antibiotics are made of gentamicin Su and kanamycin are composed according to the ratio of parts by weight of 5:2.
进一步的，所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20-40份、赖氨四环素10-30份、吉米沙星20-50份。支原体抑制剂中包括柱晶白霉素、赖氨四环素和吉米沙星能够有效抑制并杀死保护液中的支原体，作用周期短，不影响胎盘干细胞和脐带间充质干细胞的保存和代谢，并且经过处理后的细胞表面光滑，胎盘干细胞和脐带间充质干细胞在运输和后期存储过程中不会轻易被支原体重新感染，为细胞的存储提供了安全的生存环境，为胎盘或脐带的运输提供了保护作用。Further, the mycoplasma inhibitor includes the following components in parts by weight: 20-40 parts of leucomycin, 10-30 parts of lysine tetracycline, and 20-50 parts of gemifloxacin. Mycoplasma inhibitors include leucomycin, lysine tetracycline and gemifloxacin, which can effectively inhibit and kill mycoplasma in the protection solution, have a short action period, and do not affect the preservation and metabolism of placental stem cells and umbilical cord mesenchymal stem cells, and The surface of the treated cells is smooth, and the placental stem cells and umbilical cord mesenchymal stem cells will not be easily re-infected by mycoplasma during transportation and later storage, providing a safe living environment for the storage of cells and providing a safe environment for the transportation of placenta or umbilical cord. Protective effects.
进一步的，所述支原体抑制剂还包括以下重量份数的组分：莪术提取物5-10份、鱼腥草提取物4-8份、土茯苓提取物3-6份。支原体对细胞的存储有很大的威胁作用，很容易造成细胞感染，从而使细胞凋亡，为此，本发明中进一步提供了支原体抑制剂中增加了莪术提取物、鱼腥草提取物和土茯苓提取物，这三种中药成分能够在原始西药成分的基础上，通过中西医结合的药效成分组成支原体抑制剂，有效抑制和杀死保护液中的支原体，作用周期较短，对细胞具有较强的保护作用，防止细胞感染。Further, the mycoplasma inhibitor also includes the following components in parts by weight: 5-10 parts of curcuma extract, 4-8 parts of houttuynia extract, and 3-6 parts of smilax extract. Mycoplasma has very big menacing effect to the storage of cell, is easy to cause cell infection, thereby makes cell apoptosis, for this reason, the present invention further provides in the mycoplasma inhibitor that has increased turmeric extract, Houttuynia cordata extract and soil Poria cocos extract, these three Chinese medicine ingredients can form mycoplasma inhibitors through the effective ingredients of traditional Chinese and Western medicine on the basis of the original western medicine ingredients, effectively inhibit and kill mycoplasma in the protection solution, the action cycle is short, and has a certain effect on cells. Strong protective effect against cell infection.
进一步的，所述保护液还含有羟乙基淀粉和甲壳质，每mL所述MEM–α培养基水溶液分别溶解所述羟乙基淀粉8-10mg、所述甲壳质1-5mg。本发明中通过增加羟乙基淀粉和甲壳质能够有效维持胎盘干细胞和脐带间充质干细胞的渗透平衡，起到维持溶液的渗透压的作用，提高溶液稳定性，防止细胞凋亡，有效模拟人体环境，为细胞提供一个稳压、稳定、适宜的生存环境，延长了细胞的保存时间。Further, the protective solution also contains hydroxyethyl starch and chitin, 8-10 mg of the hydroxyethyl starch and 1-5 mg of the chitin are dissolved in each mL of the MEM-α medium aqueous solution. In the present invention, by adding hydroxyethyl starch and chitin, the osmotic balance of placental stem cells and umbilical cord mesenchymal stem cells can be effectively maintained, the osmotic pressure of the solution can be maintained, the stability of the solution can be improved, cell apoptosis can be prevented, and the human body can be effectively simulated. The environment provides a stable, stable and suitable living environment for cells, and prolongs the storage time of cells.
进一步的，所述保护液还含有右旋糖酐和肝素钠，每mL所述MEM–α培养基水溶液分别溶解所述右旋糖酐4-6mg、所述肝素钠1-3mg。本发明中提供的右旋糖酐和肝素钠可以作为低温保护剂，提高溶液的稳定性，为细胞提供一个与人体较为接近的生存环境，降低细胞死亡率，延长细胞的保存时间。Further, the protection solution also contains dextran and heparin sodium, and 4-6 mg of the dextran and 1-3 mg of the heparin sodium are dissolved in each mL of the MEM-α medium aqueous solution. The dextran and heparin sodium provided in the present invention can be used as cryoprotectants to improve the stability of the solution, provide cells with a living environment closer to the human body, reduce cell death rates, and prolong the storage time of cells.
进一步的，所述保护液还含有亚硒酸钠和N-乙酰半胱氨酸，每mL所述MEM–α培养基水溶液分别溶解所述亚硒酸钠0.4-0.6mg、所述N-乙酰半胱氨酸0.1-0.3mg。本发明提供的保护液中加入亚硒酸钠和N-乙酰半胱氨酸能够消除保存能引起细胞活性降低或者死亡的细胞毒素。其中，亚硒酸钠可以促进氢过氧化物代谢，消除保护液中氧化物酶和氧自由基对细胞的伤害，这一点上对干细胞具有良好的损伤保护作用。此外，N-乙酰半胱氨酸是活性氧自由基的清除剂，它可以改善氧化应激引起的不同种类细胞的损伤；对亚硒酸钠的所减缓之外的其他的氧化损伤等进行弥补，保证细胞温和的保存环境。Further, the protection solution also contains sodium selenite and N-acetyl cysteine, and each mL of the MEM-α medium aqueous solution dissolves 0.4-0.6 mg of the sodium selenite and the N-acetyl cysteine respectively. Cysteine 0.1-0.3mg. Adding sodium selenite and N-acetylcysteine to the protection solution provided by the invention can eliminate and preserve cytotoxins that can cause cell activity reduction or death. Among them, sodium selenite can promote the metabolism of hydroperoxide and eliminate the damage to cells caused by oxidase and oxygen free radicals in the protection solution, which has a good damage protection effect on stem cells. In addition, N-acetylcysteine is a scavenger of active oxygen free radicals, which can improve the damage of different types of cells caused by oxidative stress; it can compensate for other oxidative damages other than those slowed down by sodium selenite , to ensure a mild storage environment for cells.
进一步的，所述保护液还含有乳糖醛酸、血白蛋白、葡萄糖酸钠和葡萄糖胺，每mL所述MEM–α培养基水溶液分别溶解所述乳糖醛酸5-8mg、所述血白蛋白2-4mg、所述葡萄糖酸钠5-8mg、所述葡萄糖胺4-6mg。本发明中提供的保护液中加入乳糖醛酸、血白蛋白、葡萄糖酸钠、葡萄糖胺，能够为细胞提供一定的营养成分，同时能够提高细胞免疫性，防止细胞感染，抑制细胞凋亡，提高细胞的存活率。Further, the protective solution also contains lacturonic acid, serum albumin, sodium gluconate and glucosamine, and each mL of the MEM-α medium aqueous solution dissolves 5-8 mg of the lacturonic acid and the serum albumin respectively. 2-4mg, the sodium gluconate 5-8mg, the glucosamine 4-6mg. Adding lacturonic acid, serum albumin, sodium gluconate, and glucosamine to the protective solution provided in the present invention can provide certain nutrients for cells, and at the same time can improve cell immunity, prevent cell infection, inhibit cell apoptosis, and improve cell viability.
优选的，所述保护液还含有头孢哌酮钠和酚红，每mL所述MEM–α培养基水溶液分别溶解所述头孢哌酮钠1-4mg、所述酚红1-3mg。本发明中通过在保护液中加入头孢哌酮钠和酚红的作用是对可能发生的污染进行预防和提示已经发生的污染，当溶液被污染后，可以显示颜色，通过观察保护液的颜色可以判断细胞污染程度，有效减少细胞凋亡。Preferably, the protective solution also contains cefoperazone sodium and phenol red, 1-4 mg of cefoperazone sodium and 1-3 mg of phenol red are dissolved in each mL of the MEM-α medium aqueous solution. In the present invention, the effect of adding cefoperazone sodium and phenol red in the protective solution is to prevent possible pollution and prompt the pollution that has occurred. When the solution is polluted, it can display color, and the cell pollution can be judged by observing the color of the protective solution. to effectively reduce cell apoptosis.
本发明还提供了一种胎盘及脐带细胞保护液的制备方法，该制备方法包括以下步骤:The present invention also provides a kind of preparation method of placenta and umbilical cord cell protection liquid, and this preparation method comprises the following steps:
S1、取浓度为10-15mg/mL的MEM-α培养基水溶液，向所述MEM-α培养基水溶液中加入氨基糖苷类抗生素，并进行搅拌，使氨基糖苷类抗生素充分溶解在MEM-α培养基水溶液中，每mL MEM-α培养基水溶液中溶解所述氨基糖苷类抗生素的含量为180-240U；S1. Take the MEM-α medium aqueous solution with a concentration of 10-15mg/mL, add aminoglycoside antibiotics to the MEM-α medium aqueous solution, and stir, so that the aminoglycoside antibiotics are fully dissolved in the MEM-α culture In the basic aqueous solution, the content of dissolving the aminoglycoside antibiotics in every mL MEM-α medium aqueous solution is 180-240U;
S2、向步骤S1中溶解有氨基糖苷类抗生素的MEM-α培养基水溶液中加入支原体抑制剂，并进行搅拌，使支原体抑制剂充分溶解在MEM-α培养基水溶液中，即得到胎盘及脐带细胞保护液，每mL MEM-α培养基水溶液中溶解所述支原体抑制剂的含量为10-30mg。S2, add the mycoplasma inhibitor to the MEM-α medium aqueous solution that is dissolved with aminoglycoside antibiotics in step S1, and stir, make the mycoplasma inhibitor fully dissolve in the MEM-α medium aqueous solution, namely obtain placenta and umbilical cord cells For the protection solution, the content of the mycoplasma inhibitor dissolved in every mL of MEM-α medium aqueous solution is 10-30mg.
本发明与现有专利提供的保护液的制备方法相比，配制较为方便，无需特定的环境，可以大批量生产配制，不受外界环境影响。Compared with the preparation method of the protective solution provided by the existing patent, the present invention is more convenient to prepare, does not need a specific environment, can be produced and prepared in large quantities, and is not affected by the external environment.
本发明的有益效果如下：本发明提供的保护液不仅能够有效杀死或抑制支原体的形成，防止胎盘干细胞和脐带间充质干细胞感染，而且通过加入保护成分能够为胎盘干细胞和脐带间充质干细胞提供一个稳定的渗透压环境，有效模拟人体环境，提高细胞生存率，同时能够为细胞短时间内保存和运输提供营养成分，提高细胞的免疫性能，防止细胞感染，有效提高细胞的生存力，延长细胞运输时间。The beneficial effects of the present invention are as follows: the protective solution provided by the present invention can not only effectively kill or inhibit the formation of mycoplasma, prevent infection of placental stem cells and umbilical cord mesenchymal stem cells, but also protect placental stem cells and umbilical cord mesenchymal stem cells by adding protective ingredients. Provide a stable osmotic pressure environment, effectively simulate the human body environment, improve the survival rate of cells, and at the same time provide nutrients for the storage and transportation of cells in a short time, improve the immune performance of cells, prevent cell infection, effectively improve cell viability, prolong Cell transit time.
具体实施方式detailed description
下面结合以下实施例对本发明作进一步详细说明。The present invention will be described in further detail below in conjunction with the following examples.
实施例1Example 1
本发明实施例1提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂和庆大霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素180U，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 1 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors and gentamycin in MEM-α medium aqueous solution, and each mL of the MEM-α culture 10 mg of the mycoplasma inhibitor and 180 U of the gentamycin were dissolved in the base aqueous solution, and the concentration of the MEM-α medium aqueous solution was 10 mg/mL.
实施例2Example 2
本发明实施例2提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素和卡那霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素150U、所述卡那霉素60U，所述MEM–α培养基水溶液的浓度为12.5mg/mL。Embodiment 2 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors, gentamycin and kanamycin with MEM-α medium aqueous solution. The MEM-alpha medium aqueous solution dissolves 20 mg of the mycoplasma inhibitor, 150 U of the gentamycin, and 60 U of the kanamycin respectively, and the concentration of the MEM-alpha medium aqueous solution is 12.5 mg/mL.
实施例3Example 3
本发明实施例3提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、阿米卡星和妥布霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂30mg、所述庆大霉素80U、所述阿米卡星80U、所述妥布霉素80U，所述MEM–α培养基水溶液的浓度为15mg/mL。Embodiment 3 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly prepared by dissolving mycoplasma inhibitors, gentamicin, amikacin and tobramycin in an aqueous solution of MEM-α medium. 30 mg of the mycoplasma inhibitor, 80 U of the gentamycin, 80 U of the amikacin, 80 U of the tobramycin, and 80 U of the tobramycin were dissolved in each mL of the MEM-α medium aqueous solution, and the MEM-α The concentration of the aqueous medium solution was 15 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份。The mycoplasma inhibitor comprises the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, and 20 parts of gemifloxacin.
本发明还提供了一种胎盘及脐带细胞保护液的制备方法，该制备方法包括以下步骤:The present invention also provides a kind of preparation method of placenta and umbilical cord cell protection liquid, and this preparation method comprises the following steps:
S1、取浓度为15mg/mL的MEM-α培养基水溶液，向所述MEM-α培养基水溶液中加入庆大霉素、阿米卡星和妥布霉素，并进行搅拌，使庆大霉素、阿米卡星和妥布霉素充分溶解在MEM-α培养基水溶液中，每mL MEM-α培养基水溶液中溶解所述庆大霉素80U、所述阿米卡星80U、所述妥布霉素80U；S1, get the MEM-α medium aqueous solution that concentration is 15mg/mL, add gentamicin, amikacin and tobramycin in described MEM-α medium aqueous solution, and stir, make gentamicin Sufficiently dissolve gentamycin, amikacin and tobramycin in MEM-α medium aqueous solution, dissolve described gentamycin 80U, described amikacin 80U, described Tobramycin 80U;
S2、向步骤S1中溶解有庆大霉素、阿米卡星和妥布霉素的MEM-α培养基水溶液中加入支原体抑制剂，并进行搅拌，使支原体抑制剂充分溶解在MEM-α培养基水溶液中，即得到胎盘及脐带细胞保护液，每mL MEM-α培养基水溶液中溶解所述支原体抑制剂的含量为30mg。S2, add mycoplasma inhibitor to the MEM-α culture medium aqueous solution that has gentamicin, amikacin and tobramycin dissolved in step S1, and stir, make mycoplasma inhibitor fully dissolve in MEM-α culture In the aqueous base solution, the placenta and umbilical cord cell protection solution is obtained, and the content of the mycoplasma inhibitor dissolved in each mL of MEM-α medium aqueous solution is 30 mg.
支原体抑制剂的配制方法为：将柱晶白霉素20份、赖氨四环素10份、吉米沙星20份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: directly mix 20 parts of leucomycin, 10 parts of lysine tetracycline and 20 parts of gemifloxacin.
实施例4Example 4
本发明实施例4提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、卡那霉素和阿米卡星用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素70U、所述卡那霉素70U和所述阿米卡星70U，所述MEM–α培养基水溶液的浓度为11mg/mL。Embodiment 4 of the present invention provides a placenta and umbilical cord cell protection solution, the protection solution is mainly prepared by dissolving mycoplasma inhibitor, gentamicin, kanamycin and amikacin with MEM-α medium aqueous solution 20 mg of the mycoplasma inhibitor, the 70 U of the gentamycin, the 70 U of the kanamycin and the 70 U of the amikacin were dissolved in each mL of the MEM-α medium aqueous solution, and the MEM-α The concentration of the aqueous medium solution was 11 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素30份、赖氨四环素20份、吉米沙星35份。The mycoplasma inhibitor comprises the following components in parts by weight: 30 parts of leucomycin, 20 parts of lysine tetracycline, and 35 parts of gemifloxacin.
本实施例4中提供的保护液的制备方法与实施例3的方法相同。The preparation method of the protective solution provided in Example 4 is the same as that in Example 3.
支原体抑制剂的配制方法为：将柱晶白霉素30份、赖氨四环素20份、吉米沙星35份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: directly mix 30 parts of leucomycin, 20 parts of lysine tetracycline and 35 parts of gemifloxacin.
实施例5Example 5
本发明实施例5提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、卡那霉素、阿米卡星和妥布霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素60U、所述卡那霉素60U、所述阿米卡星30U、所述妥布霉素30U，所述MEM–α培养基水溶液的浓度为12mg/mL。Embodiment 5 of the present invention provides a kind of placenta and umbilical cord cell protection liquid, and described protection liquid mainly is mycoplasma inhibitor, gentamicin, kanamycin, amikacin and tobramycin with MEM-α The aqueous medium solution is dissolved, and each mL of the MEM-α medium aqueous solution dissolves the mycoplasma inhibitor 10mg, the gentamicin 60U, the kanamycin 60U, the amikacin 30U, The tobramycin is 30 U, and the concentration of the MEM-α medium aqueous solution is 12 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份。The mycoplasma inhibitor includes the following components in parts by weight: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of houttuynia extract, Smilax smilax Extract 3 parts.
本实施例5中提供的保护液的制备方法与实施例3的方法相同。The preparation method of the protective solution provided in Example 5 is the same as that in Example 3.
支原体抑制剂的配制方法为：将柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of Houttuynia cordata extract, 3 parts of Smilax tuckahoe extract Just mix it.
实施例6Example 6
本发明实施例6提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、卡那霉素、阿米卡星和妥布霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素60U、所述卡那霉素60U、所述阿米卡星30U、所述妥布霉素30U，所述MEM–α培养基水溶液的浓度为12mg/mL。Embodiment 6 of the present invention provides a placenta and umbilical cord cell protection solution, the protection solution is mainly mycoplasma inhibitors, gentamicin, kanamycin, amikacin and tobramycin with MEM-α The aqueous medium solution is dissolved, and each mL of the MEM-α medium aqueous solution dissolves the mycoplasma inhibitor 10mg, the gentamicin 60U, the kanamycin 60U, the amikacin 30U, The tobramycin is 30 U, and the concentration of the MEM-α medium aqueous solution is 12 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份。The mycoplasma inhibitor includes the following components in parts by weight: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of zedoary extract, 6 parts of houttuynia extract, Smilax smilax Extract 5 parts.
本实施例6中提供的保护液的制备方法与实施例3的方法相同。The preparation method of the protective solution provided in Example 6 is the same as that in Example 3.
支原体抑制剂的配制方法为：将柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份直接混合即可。The preparation method of mycoplasma inhibitor is as follows: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of curcuma extract, 6 parts of Houttuynia cordata extract, and 5 parts of Smilax cocos extract are directly Just mix it.
实施例7Example 7
本发明实施例7提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉和甲壳质用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉10mg和所述甲壳质1mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 7 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors, gentamicin, hydroxyethyl starch and chitin in an aqueous MEM-α medium solution. Every mL of the MEM-alpha medium aqueous solution dissolves respectively the mycoplasma inhibitor 10mg, the gentamycin 180U, the hydroxyethyl starch 10mg and the chitin 1mg, the MEM-alpha medium aqueous solution The concentration is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份。The mycoplasma inhibitor includes the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of houttuynia extract, Extract 6 parts.
本实施例7提供的保护液的制备方法是将支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉10mg和所述甲壳质1mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 7 is to dissolve 10 mg of mycoplasma inhibitor, 180 U of gentamycin, 10 mg of hydroxyethyl starch and 1 mg of chitin respectively in an aqueous solution of MEM-alpha medium. Can.
支原体抑制剂的配制方法为：将柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of Houttuynia cordata extract, and 6 parts of Smilax tuckahoe extract Just mix it.
实施例8Example 8
本发明实施例8提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉和甲壳质用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉8mg和所述甲壳质3mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 8 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly formed by dissolving mycoplasma inhibitors, gentamicin, hydroxyethyl starch and chitin in an aqueous solution of MEM-α medium. Every mL of the MEM-alpha medium aqueous solution dissolves respectively the mycoplasma inhibitor 10mg, the gentamycin 180U, the hydroxyethyl starch 8mg and the chitin 3mg, the MEM-alpha medium aqueous solution The concentration is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份。The mycoplasma inhibitor includes the following components in parts by weight: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of houttuynia extract, Smilax smilax Extract 3 parts.
本实施例8提供的保护液的制备方法是将支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉8mg和所述甲壳质3mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 8 is to dissolve 10 mg of mycoplasma inhibitor, 180 U of gentamycin, 8 mg of hydroxyethyl starch and 3 mg of chitin in MEM-alpha medium aqueous solution respectively. Can.
支原体抑制剂的配制方法为：将柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of Houttuynia cordata extract, 3 parts of Smilax tuckahoe extract Just mix it.
实施例9Example 9
本发明实施例9提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐和肝素钠用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素210U、所述羟乙基淀粉9mg、所述甲壳质5mg、所述右旋糖酐4mg、所述肝素钠1mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 9 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly composed of mycoplasma inhibitors, gentamicin, hydroxyethyl starch, chitin, dextran and heparin sodium in MEM-α medium Each mL of the MEM-α medium aqueous solution dissolves the mycoplasma inhibitor 20mg, the gentamicin 210U, the hydroxyethyl starch 9mg, the chitin 5mg, and the dextran 4mg 1. The sodium heparin is 1 mg, and the concentration of the MEM-α medium aqueous solution is 10 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份。The mycoplasma inhibitor includes the following components in parts by weight: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of zedoary extract, 6 parts of houttuynia extract, Smilax smilax Extract 5 parts.
本实施例9提供的保护液的制备方法是将支原体抑制剂20mg、庆大霉素210U、羟乙基淀粉9mg、甲壳质5mg、右旋糖酐4mg、肝素钠1mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 9 is to dissolve 20 mg of mycoplasma inhibitor, 210 U of gentamicin, 9 mg of hydroxyethyl starch, 5 mg of chitin, 4 mg of dextran, and 1 mg of sodium heparin in the aqueous solution of MEM-α medium That's it.
支原体抑制剂的配制方法为：将柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份直接混合即可。The preparation method of mycoplasma inhibitor is as follows: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of curcuma extract, 6 parts of Houttuynia cordata extract, and 5 parts of Smilax cocos extract are directly Just mix it.
实施例10Example 10
本发明实施例10提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠、亚硒酸钠和N-乙酰半胱氨酸用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉8mg、所述甲壳质3mg、所述右旋糖酐6mg、所述肝素钠3mg、所述亚硒酸钠0.4mg、所述N-乙酰半胱氨酸0.1mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 10 of the present invention provides a kind of placenta and umbilical cord cell protection liquid, and described protection liquid mainly is mycoplasma inhibitor, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium, sodium selenite and N-acetylcysteine is formed by dissolving MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution dissolves 10 mg of the mycoplasma inhibitor, 180 U of the gentamicin, and 180 U of the hydroxyethyl 8 mg of starch, 3 mg of chitin, 6 mg of dextran, 3 mg of sodium heparin, 0.4 mg of sodium selenite, 0.1 mg of N-acetylcysteine, and the aqueous solution of MEM-α medium The concentration is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份。The mycoplasma inhibitor includes the following components in parts by weight: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of houttuynia extract, Smilax smilax Extract 3 parts.
本实施例10提供的保护液的制备方法是将支原体抑制剂10mg、庆大霉素180U、羟乙基淀粉8mg、甲壳质3mg、右旋糖酐6mg、肝素钠3mg、亚硒酸钠0.4mg、N-乙酰半胱氨酸0.1mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 10 is to mix mycoplasma inhibitor 10mg, gentamicin 180U, hydroxyethyl starch 8mg, chitin 3mg, dextran 6mg, heparin sodium 3mg, sodium selenite 0.4mg, N- Acetylcysteine 0.1 mg was dissolved in MEM-α medium aqueous solution.
支原体抑制剂的配制方法为：将柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of Houttuynia cordata extract, 3 parts of Smilax tuckahoe extract Just mix it.
实施例11Example 11
本发明实施例11提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠、亚硒酸钠、N-乙酰半胱氨酸、乳糖醛酸、血白蛋白、葡萄糖酸钠和葡萄糖胺用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素210U、所述羟乙基淀粉9mg、所述甲壳质1mg、所述右旋糖酐4mg、所述肝素钠1mg、所述亚硒酸钠0.6mg、所述N-乙酰半胱氨酸0.3mg、所述乳糖醛酸5mg、所述血白蛋白2mg、所述葡萄糖酸钠5mg、所述葡萄糖胺4mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 11 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly composed of mycoplasma inhibitors, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium, sodium selenite, N-acetyl cysteine, lacturonic acid, serum albumin, sodium gluconate and glucosamine are formed by dissolving MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution dissolves the mycoplasma inhibitor respectively 20mg, the gentamicin 210U, the hydroxyethyl starch 9mg, the chitin 1mg, the dextran 4mg, the heparin sodium 1mg, the sodium selenite 0.6mg, the N-acetyl semi Cystine 0.3 mg, lacturonic acid 5 mg, serum albumin 2 mg, sodium gluconate 5 mg, glucosamine 4 mg, and the concentration of the MEM-α medium aqueous solution was 10 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份。The mycoplasma inhibitor includes the following components in parts by weight: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of zedoary extract, 6 parts of houttuynia extract, Smilax smilax Extract 5 parts.
本实施例11提供的保护液的制备方法是将支原体抑制剂20mg、庆大霉素210U、羟乙基淀粉9mg、甲壳质1mg、右旋糖酐4mg、肝素钠1mg、亚硒酸钠0.6mg、N-乙酰半胱氨酸0.3mg、乳糖醛酸5mg、血白蛋白2mg、葡萄糖酸钠5mg、葡萄糖胺4mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in this example 11 is to mix mycoplasma inhibitor 20mg, gentamicin 210U, hydroxyethyl starch 9mg, chitin 1mg, dextran 4mg, heparin sodium 1mg, sodium selenite 0.6mg, N- Acetyl cysteine 0.3 mg, lacturonic acid 5 mg, serum albumin 2 mg, sodium gluconate 5 mg, glucosamine 4 mg were dissolved in MEM-α medium aqueous solution.
支原体抑制剂的配制方法为：将柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份直接混合即可。The preparation method of mycoplasma inhibitor is as follows: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of curcuma extract, 6 parts of Houttuynia cordata extract, and 5 parts of Smilax cocos extract are directly Just mix it.
实施例12Example 12
本发明实施例12提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠、亚硒酸钠、N-乙酰半胱氨酸、乳糖醛酸、血白蛋白、葡萄糖酸钠、葡萄糖胺、头孢哌酮钠和酚红用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂10mg、所述庆大霉素180U、所述羟乙基淀粉8mg、所述甲壳质3mg、所述右旋糖酐6mg、所述肝素钠3mg、所述亚硒酸钠0.4mg、所述N-乙酰半胱氨酸0.1mg、所述乳糖醛酸8mg、所述血白蛋白4mg、所述葡萄糖酸钠8mg、所述葡萄糖胺6mg、所述头孢哌酮钠1mg、所述酚红1mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 12 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly composed of mycoplasma inhibitors, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium, sodium selenite, N-acetylcysteine, lacturonic acid, serum albumin, sodium gluconate, glucosamine, cefoperazone sodium and phenol red are dissolved in MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is dissolved separately The mycoplasma inhibitor 10mg, the gentamicin 180U, the hydroxyethyl starch 8mg, the chitin 3mg, the dextran 6mg, the heparin sodium 3mg, the sodium selenite 0.4mg, the 0.1 mg of N-acetylcysteine, 8 mg of lacturonic acid, 4 mg of serum albumin, 8 mg of sodium gluconate, 6 mg of glucosamine, 1 mg of cefoperazone sodium, and 1 mg of phenol red. The concentration of the MEM-α medium aqueous solution is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份。The mycoplasma inhibitor includes the following components in parts by weight: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of houttuynia extract, Smilax smilax Extract 3 parts.
本实施例12提供的保护液的制备方法是将支原体抑制剂10mg、庆大霉素180U、羟乙基淀粉8mg、甲壳质3mg、右旋糖酐6mg、肝素钠3mg、亚硒酸钠0.4mg、N-乙酰半胱氨酸0.1mg、乳糖醛酸8mg、血白蛋白4mg、葡萄糖酸钠8mg、葡萄糖胺6mg、头孢哌酮钠1mg、酚红1mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 12 is to mix mycoplasma inhibitor 10mg, gentamicin 180U, hydroxyethyl starch 8mg, chitin 3mg, dextran 6mg, heparin sodium 3mg, sodium selenite 0.4mg, N- Acetylcysteine 0.1 mg, lacturonic acid 8 mg, serum albumin 4 mg, sodium gluconate 8 mg, glucosamine 6 mg, cefoperazone sodium 1 mg, and phenol red 1 mg were dissolved in MEM-α medium aqueous solution.
支原体抑制剂的配制方法为：将柱晶白霉素40份、赖氨四环素30份、吉米沙星50份、莪术提取物5份、鱼腥草提取物4份、土茯苓提取物3份直接混合即可。The preparation method of the mycoplasma inhibitor is as follows: 40 parts of leucomycin, 30 parts of lysine tetracycline, 50 parts of gemifloxacin, 5 parts of zedoary extract, 4 parts of Houttuynia cordata extract, 3 parts of Smilax tuckahoe extract Just mix it.
实施例13Example 13
本发明实施例11提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠、亚硒酸钠、N-乙酰半胱氨酸、乳糖醛酸、血白蛋白、葡萄糖酸钠、葡萄糖胺、头孢哌酮钠和酚红用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素210U、所述羟乙基淀粉9mg、所述甲壳质4mg、所述右旋糖酐4mg、所述肝素钠1mg、所述亚硒酸钠0.6mg、所述N-乙酰半胱氨酸0.3mg、所述乳糖醛酸5mg、所述血白蛋白2mg、所述葡萄糖酸钠5mg、所述葡萄糖胺4mg、所述头孢哌酮钠4mg、所述酚红3mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Embodiment 11 of the present invention provides a protective solution for placenta and umbilical cord cells. The protective solution is mainly composed of mycoplasma inhibitors, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium, sodium selenite, N-acetylcysteine, lacturonic acid, serum albumin, sodium gluconate, glucosamine, cefoperazone sodium and phenol red are dissolved in MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is dissolved separately The mycoplasma inhibitor 20mg, the gentamicin 210U, the hydroxyethyl starch 9mg, the chitin 4mg, the dextran 4mg, the heparin sodium 1mg, the sodium selenite 0.6mg, the 0.3 mg of N-acetylcysteine, 5 mg of lacturonic acid, 2 mg of serum albumin, 5 mg of sodium gluconate, 4 mg of glucosamine, 4 mg of cefoperazone sodium, and 3 mg of phenol red. The concentration of the MEM-α medium aqueous solution is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份。The mycoplasma inhibitor includes the following components in parts by weight: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of zedoary extract, 6 parts of houttuynia extract, Smilax smilax Extract 5 parts.
本实施例13提供的保护液的制备方法是将支原体抑制剂20mg、庆大霉素210U、羟乙基淀粉9mg、甲壳质4mg、右旋糖酐4mg、肝素钠1mg、亚硒酸钠0.6mg、N-乙酰半胱氨酸0.3mg、乳糖醛酸5mg、血白蛋白2mg、葡萄糖酸钠5mg、葡萄糖胺4mg、头孢哌酮钠4mg、酚红3mg分别溶解在MEM–α培养基水溶液中即可。The preparation method of the protective solution provided in Example 13 is to mix mycoplasma inhibitor 20mg, gentamicin 210U, hydroxyethyl starch 9mg, chitin 4mg, dextran 4mg, heparin sodium 1mg, sodium selenite 0.6mg, N- Acetylcysteine 0.3 mg, lacturonic acid 5 mg, serum albumin 2 mg, sodium gluconate 5 mg, glucosamine 4 mg, cefoperazone sodium 4 mg, and phenol red 3 mg were dissolved in MEM-α medium aqueous solution.
支原体抑制剂的配制方法为：将柱晶白霉素30份、赖氨四环素20份、吉米沙星35份、莪术提取物8份、鱼腥草提取物6份、土茯苓提取物5份直接混合即可。The preparation method of mycoplasma inhibitor is as follows: 30 parts of leucomycin, 20 parts of lysine tetracycline, 35 parts of gemifloxacin, 8 parts of curcuma extract, 6 parts of Houttuynia cordata extract, and 5 parts of Smilax cocos extract are directly Just mix it.
对照例1Comparative example 1
本发明对照例1提供了一种胎盘及脐带细胞保护液，所述保护液主要是将庆大霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述庆大霉素100U，所述MEM–α培养基水溶液的浓度为10mg/mL。Comparative Example 1 of the present invention provides a protective solution for placenta and umbilical cord cells, the protective solution is mainly formed by dissolving gentamicin with MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is dissolved respectively The gentamicin is 100 U, and the concentration of the MEM-α medium aqueous solution is 10 mg/mL.
对照例2Comparative example 2
本发明对照例2提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg，所述MEM–α培养基水溶液的浓度为10mg/mL。Comparative Example 2 of the present invention provides a protective solution for placenta and umbilical cord cells, the protective solution is mainly formed by dissolving mycoplasma inhibitors with MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution is dissolved respectively The mycoplasma inhibitor 20mg, the concentration of the MEM-α medium aqueous solution is 10mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素15份。The mycoplasma inhibitor comprises the following components in parts by weight: 15 parts of leucomycin.
对照例3Comparative example 3
本发明对照例3提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂和庆大霉素用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素180U，所述MEM–α培养基水溶液的浓度为10mg/mL。Comparative Example 3 of the present invention provides a protective solution for placenta and umbilical cord cells, the protective solution is mainly formed by dissolving mycoplasma inhibitors and gentamycin with MEM-α medium aqueous solution, and each mL of MEM-α culture 20 mg of the mycoplasma inhibitor and 180 U of the gentamycin were dissolved in the base aqueous solution, and the concentration of the MEM-α medium aqueous solution was 10 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：赖氨四环素30份、莪术提取物5份、土茯苓提取物3份。The mycoplasma inhibitor includes the following components in parts by weight: 30 parts of lysine tetracycline, 5 parts of curcuma extract, and 3 parts of Smilax smilax extract.
对照例4Comparative example 4
本发明对照例4提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素和羟乙基淀粉用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、所述庆大霉素180U、所述羟乙基淀粉10mg，所述MEM–α培养基水溶液的浓度为12.5mg/mL。Comparative example 4 of the present invention provides a kind of placenta and umbilical cord cell protective solution, and described protective solution is mainly that mycoplasma inhibitor, gentamicin and hydroxyethyl starch are dissolved with MEM-alpha culture medium aqueous solution, and each mL The MEM-alpha medium aqueous solution dissolves 20 mg of the mycoplasma inhibitor, 180 U of the gentamycin, and 10 mg of the hydroxyethyl starch respectively, and the concentration of the MEM-alpha medium aqueous solution is 12.5 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份。The mycoplasma inhibitor includes the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of houttuynia extract, Extract 6 parts.
对照例5Comparative example 5
本发明对照例5提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质和右旋糖酐用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、庆大霉素180U、羟乙基淀粉9mg、甲壳质1mg、右旋糖酐4mg，所述MEM–α培养基水溶液的浓度为12.5mg/mL。Comparative example 5 of the present invention provides a kind of placenta and umbilical cord cell protective solution, and described protective solution is mainly that mycoplasma inhibitor, gentamicin, hydroxyethyl starch, chitin and dextran are dissolved with MEM-alpha culture medium aqueous solution. 20 mg of the mycoplasma inhibitor, 180 U of gentamicin, 9 mg of hydroxyethyl starch, 1 mg of chitin, and 4 mg of dextran were dissolved in each mL of the MEM-α medium aqueous solution, and the concentration of the MEM-α medium aqueous solution was It is 12.5mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份。The mycoplasma inhibitor includes the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of houttuynia extract, Extract 6 parts.
对照例6Comparative example 6
本发明对照例6提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠和亚硒酸钠用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、庆大霉素180U、羟乙基淀粉9mg、甲壳质4mg、右旋糖酐4mg、肝素钠3mg、亚硒酸钠0.4mg，所述MEM–α培养基水溶液的浓度为14mg/mL。Comparative example 6 of the present invention provides a kind of placenta and umbilical cord cell protection solution, and described protection solution mainly is mycoplasma inhibitor, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium and sodium selenite MEM-α medium aqueous solution is dissolved, and each mL of the MEM-α medium aqueous solution dissolves 20 mg of the mycoplasma inhibitor, 180 U of gentamicin, 9 mg of hydroxyethyl starch, 4 mg of chitin, 4 mg of dextran, and sodium heparin 3mg, sodium selenite 0.4mg, the concentration of the MEM-α medium aqueous solution is 14mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份。The mycoplasma inhibitor includes the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of houttuynia extract, Extract 6 parts.
对照例7Comparative example 7
本发明对照例6提供了一种胎盘及脐带细胞保护液，所述保护液主要是将支原体抑制剂、庆大霉素、羟乙基淀粉、甲壳质、右旋糖酐、肝素钠、亚硒酸钠、N-乙酰半胱氨酸、乳糖醛酸和血白蛋白用MEM–α培养基水溶液溶解而成，每mL所述MEM–α培养基水溶液分别溶解所述支原体抑制剂20mg、庆大霉素180U、羟乙基淀粉9mg、甲壳质5mg、右旋糖酐4mg、肝素钠3mg、亚硒酸钠0.4mg、N-乙酰半胱氨酸0.3mg、乳糖醛酸5mg、血白蛋白2mg，所述MEM–α培养基水溶液的浓度为14mg/mL。Comparative example 6 of the present invention provides a kind of placenta and umbilical cord cell protection liquid, and described protection liquid mainly is mycoplasma inhibitor, gentamicin, hydroxyethyl starch, chitin, dextran, heparin sodium, sodium selenite, N-acetylcysteine, lacturonic acid and serum albumin are formed by dissolving MEM-α medium aqueous solution, and each mL of the MEM-α medium aqueous solution dissolves 20 mg of the mycoplasma inhibitor and 180 U of gentamicin respectively , hydroxyethyl starch 9mg, chitin 5mg, dextran 4mg, heparin sodium 3mg, sodium selenite 0.4mg, N-acetylcysteine 0.3mg, lacturonic acid 5mg, serum albumin 2mg, the MEM-α The concentration of the aqueous medium solution was 14 mg/mL.
所述支原体抑制剂包括以下重量份数的组分：柱晶白霉素20份、赖氨四环素10份、吉米沙星20份、莪术提取物10份、鱼腥草提取物8份、土茯苓提取物6份。The mycoplasma inhibitor includes the following components in parts by weight: 20 parts of leucomycin, 10 parts of lysine tetracycline, 20 parts of gemifloxacin, 10 parts of zedoary extract, 8 parts of houttuynia extract, Extract 6 parts.
试验1、本发明提供的保护液对胎盘干细胞和脐带间充质干细胞存活率的影响Test 1, the protection solution provided by the present invention has the influence on placental stem cell and umbilical cord mesenchymal stem cell survival rate
1、检测样品：1. Test samples:
取本发明实施例1、实施例2和对照例1提供的保护液进行检测试验。The protective solutions provided in Example 1, Example 2 and Comparative Example 1 of the present invention were used for detection tests.
2、采集样品：2. Collect samples:
分别3组细胞群，第一组细胞群中含有1.9×105个离体胎盘干细胞和脐带间充质干细胞，第二组细胞群中含有2.5×105个离体胎盘干细胞和脐带间充质干细胞，第三组细胞群中含有2.2×105个离体胎盘干细胞和脐带间充质干细胞，将第一组、第二组和第三组细胞群分别浸没在实施例1、实施例2和对照例1提供的保护液中，在温度为4℃的条件下保存3天后，检测不同保护液中胎盘干细胞和脐带间充质干细胞的数量和生长情况。Three groups of cell populations, the first group of cell populations contained 1.9×105 isolated placental stem cells and umbilical cord mesenchymal stem cells, and the second group of cell populations contained 2.5 × 105 isolated placental stem cells and umbilical cord mesenchymal stem cells Stem cells, the third group of cell populations contains 2.2×10 5 isolated placental stem cells and umbilical cord mesenchymal stem cells, the first group, the second group and the third group of cell populations were respectively immersed in the In the protection solution provided in Control Example 1, after being stored at 4° C. for 3 days, the number and growth of placental stem cells and umbilical cord mesenchymal stem cells in different protection solutions were detected.
由上述数据得知，本发明实施例1、实施例2提供的保护液与对照例1相比，本发明实施例1、实施例2提供的保护液能够有效提高细胞存活率，而且保护液中支原体抑制剂和庆大霉素的含量越高，其细胞的存活率越高，实施例2提供的保护液中的细胞存活率达到了95％，为此，可以验证本发明提供的保护液中通过在MEM–α培养基溶液中加入支原体抑制剂和氨基糖苷类抗生素能够有效模拟人体环境，提高细胞存活率，降低细胞凋亡，延长细胞运输时间，并且实施例2加入混合的氨基糖苷类抗生素的效果明显好于单一的氨基糖苷类抗生素。From the above data, it can be known that the protective solution provided by Example 1 and Example 2 of the present invention is compared with Comparative Example 1, and the protective solution provided by Example 1 and Example 2 of the present invention can effectively improve the cell survival rate, and in the protective solution The higher the content of mycoplasma inhibitor and gentamicin, the higher the survival rate of the cells, the cell survival rate in the protection solution provided by Example 2 reached 95%, for this reason, it can be verified that in the protection solution provided by the present invention By adding mycoplasma inhibitors and aminoglycoside antibiotics to the MEM-α medium solution, the human environment can be effectively simulated, the cell survival rate can be improved, cell apoptosis can be reduced, and the cell transportation time can be prolonged, and the mixed aminoglycoside antibiotics can be added in Example 2 The effect is significantly better than a single aminoglycoside antibiotics.
试验2、本发明提供的保护液对胎盘干细胞和脐带间充质干细胞的活率检测Test 2, the protective solution provided by the present invention detects the viability of placental stem cells and umbilical cord mesenchymal stem cells
1、检测样品：以实施例1、7、9、11提供的保护液为实验1-4组，对照例1、4、5、7提供的保护液对照1-4组。1. Test samples: use the protective solutions provided in Examples 1, 7, 9 and 11 as experimental groups 1-4, and the protective solutions provided in Comparative Examples 1, 4, 5 and 7 as control groups 1-4.
2、采集样品：将采集的等量的脐带细胞或胎盘细胞分别浸没上述实验1-4组和对照1-4组中。2. Collect samples: Submerge the collected equal amount of umbilical cord cells or placental cells into the above experimental groups 1-4 and control groups 1-4 respectively.
将各实验组和对照组的脐带细胞或胎盘细胞分别在24h、48h、96h、240h、480h、960h和1920h进入试验程序，调整脐带细胞或胎盘细胞的密度均为1×106cells/mL。按细胞悬液：0.4％台盼蓝＝3:1(v:v)充分混匀，取20μL细胞悬液加入细胞计数板中，用Countstar细胞计数器进行细胞活性率检测。细胞活性率结果如下表所示。The umbilical cord cells or placental cells of each experimental group and control group were put into the test program at 24h, 48h, 96h, 240h, 480h, 960h and 1920h respectively, and the density of umbilical cord cells or placental cells was adjusted to 1×10 6 cells/mL. According to the cell suspension: 0.4% trypan blue = 3:1 (v:v), mix thoroughly, take 20 μL of the cell suspension and add it to the cell counting plate, and use the Countstar cell counter to detect the cell viability. The cell viability results are shown in the table below.
实验组和对照组细胞活率结果Cell viability results of experimental group and control group
注：“--”表示没有活性。Note: "--" indicates no activity.
由上述数据可知，实施例1提供的保护液保存的胎盘干细胞和脐带间充质干细胞的细胞活性要比使用对照例1提供的保护液保存的胎盘干细胞和脐带间充质干细胞的细胞活率高；实施例7、9、11分别提供的保护液保存的胎盘干细胞和脐带间充质干细胞的活性要比使用对照例1、4、5、7提供的保护液保存的胎盘干细胞和脐带间充质干细胞的细胞活率高；并且实施例9和11提供的保护液保存的胎盘干细胞和脐带间充质干细胞的时间要比对照例5和7提供保护液保存的胎盘干细胞和脐带间充质干细胞的时间长，分别可达到480h和1920h。It can be seen from the above data that the cell viability of the placental stem cells and umbilical cord mesenchymal stem cells preserved in the protective solution provided in Example 1 is higher than that of the placental stem cells and umbilical cord mesenchymal stem cells preserved in the protective solution provided in Comparative Example 1 The activity of the placental stem cells and umbilical cord mesenchymal stem cells preserved by the protective solution provided by Examples 7, 9 and 11 is more than that of the placental stem cells and umbilical cord mesenchymal stem cells preserved by the protective solution provided by comparative examples 1, 4, 5 and 7 The cell viability of stem cells is high; and the placental stem cells and umbilical cord mesenchymal stem cells preserved by the protective solution provided by Examples 9 and 11 provide the placental stem cells and umbilical cord mesenchymal stem cells preserved by the protective solution compared with control examples 5 and 7. The time is long, which can reach 480h and 1920h respectively.
由此得出，本发明提供的保护液中缺少支原体抑制剂或氨基糖苷类抗生素后，其干细胞的活率显著降低。当保护液中选择的MEM–α培养基溶液为加入羟乙基淀粉和甲壳质后，能够显著提高保护液保存的胎盘干细胞和脐带间充质干细胞的活率，当羟乙基淀粉和甲壳质的成分中缺少其一或者成分被替代后，失去了提高干细胞活率的作用；并且在保护液中加入右旋糖酐和肝素钠的混合物还可提高保护液胎盘干细胞和脐带间充质干细胞的保存时间，可将保存时间提高到480h，当右旋糖酐和肝素钠的成分缺少其一或被别的成分替代后，胎盘干细胞和脐带间充质干细胞的保存时间会缩短；本发明提供的保护液中加入乳糖醛酸、血白蛋白、葡萄糖酸钠和葡萄糖胺的混合物还可提高保护液胎盘干细胞和脐带间充质干细胞的保存时间，保存时间能够达到1920h，细胞的活率依然保持在65％以上。It can be concluded that the viability of stem cells is significantly reduced when the protective solution provided by the present invention lacks mycoplasma inhibitors or aminoglycoside antibiotics. When the MEM-α medium solution selected in the protection solution is added with hydroxyethyl starch and chitin, it can significantly improve the viability of placental stem cells and umbilical cord mesenchymal stem cells preserved in the protection solution. When hydroxyethyl starch and chitin If one of the components is missing or the components are replaced, the effect of increasing the viability of stem cells will be lost; and adding a mixture of dextran and sodium heparin to the protection solution can also improve the preservation time of placental stem cells and umbilical cord mesenchymal stem cells in the protection solution. The storage time can be increased to 480h. When the components of dextran and heparin sodium lack one of them or are replaced by other components, the storage time of placental stem cells and umbilical cord mesenchymal stem cells will be shortened; the protection solution provided by the present invention is added with lacturaldehyde The mixture of acid, serum albumin, sodium gluconate and glucosamine can also improve the storage time of placental stem cells and umbilical cord mesenchymal stem cells in the protective solution. The storage time can reach 1920h, and the cell viability remains above 65%.
试验3、本发明提供的保护液对胎盘干细胞和脐带间充质干细胞免疫力的影响Test 3, the protective solution provided by the present invention has an impact on the immunity of placental stem cells and umbilical cord mesenchymal stem cells
1、检测样品：1. Test samples:
取本发明实施例3、5、10和对照例2、3、6提供的保护液进行检测试验。The protective solutions provided by Examples 3, 5, and 10 of the present invention and Comparative Examples 2, 3, and 6 were used for detection tests.
2、采集样品：2. Collect samples:
分别6组细胞群，第一组细胞群中含有7.9×105个离体胎盘干细胞和脐带间充质干细胞，第二组细胞群中含有3.2×105个离体胎盘干细胞和脐带间充质干细胞，第三组细胞群中含有4.9×105个离体胎盘干细胞和脐带间充质干细胞，第四组细胞群中含有3.5×105个离体胎盘干细胞和脐带间充质干细胞，第五组细胞群中含有4.2×105个离体胎盘干细胞和脐带间充质干细胞，第六组细胞群中含有3.2×105个离体胎盘干细胞和脐带间充质干细胞，将第一、二、三、四、五、六组细胞群分别浸没在实施例3、5、10及对照例2、3、6提供的保护液中，在温度为4℃的条件下保存12天后，将培养后的胎盘干细胞和脐带间充质干细胞消化处理后用流式细胞仪计数检测细胞感染支原体或其他细菌的数量，并计算细胞受到支原体或细菌感染率；结果如下表：。There were 6 groups of cell populations, the first group of cell populations contained 7.9×10 5 isolated placental stem cells and umbilical cord mesenchymal stem cells, and the second group of cell populations contained 3.2×10 5 isolated placental stem cells and umbilical cord mesenchymal stem cells Stem cells, the third group of cell populations contained 4.9×105 isolated placental stem cells and umbilical cord mesenchymal stem cells, the fourth group of cell populations contained 3.5 ×105 isolated placental stem cells and umbilical cord mesenchymal stem cells, the fifth The first cell group contained 4.2×10 5 isolated placental stem cells and umbilical cord mesenchymal stem cells, and the sixth group cell group contained 3.2×10 5 isolated placental stem cells and umbilical cord mesenchymal stem cells. Three, four, five, and six groups of cell populations were submerged in the protection solutions provided in Examples 3, 5, 10 and Comparative Examples 2, 3, and 6 respectively, and after being stored at 4°C for 12 days, the cultured After the placental stem cells and umbilical cord mesenchymal stem cells were digested, flow cytometry was used to count the number of cells infected with mycoplasma or other bacteria, and the rate of cells infected with mycoplasma or bacteria was calculated; the results are shown in the following table:.
由上述数据得出，由实施例3和对照例2对比可知，本发明提供的保护液中含有的庆大霉素和支原体抑制剂能够有效抑制细菌或支原体的生成，同时能够杀死支原体，确认任意一种成分均不能达到抑制细菌或支原体的效果。由实施例5和对照例3可知，中西医结合组成的支原体抑制剂能够有效降低支原体和细菌的形成，而且本发明中公开了支原体抑制剂由柱晶白霉素、赖氨四环素、吉米沙星、莪术提取物、鱼腥草提取物、土茯苓提取物能够比较有效的抑制和杀死支原体，由实施例10和对照例6可知，本发明提供的保护液能够有效提高细胞的免疫能力，防止细胞感染支原体。From the above data, it can be seen from the comparison of Example 3 and Comparative Example 2 that the gentamicin and mycoplasma inhibitors contained in the protective solution provided by the present invention can effectively inhibit the generation of bacteria or mycoplasma, and can kill mycoplasma simultaneously, confirming None of the ingredients could achieve the effect of inhibiting bacteria or mycoplasma. From Example 5 and Comparative Example 3, it can be seen that the mycoplasma inhibitor formed by the combination of traditional Chinese and western medicine can effectively reduce the formation of mycoplasma and bacteria, and the present invention discloses that the mycoplasma inhibitor is composed of leumycin, lysine tetracycline, gemifloxacin , zedoary extract, Houttuynia cordata extract, Smilax tuckahoe extract can more effectively inhibit and kill mycoplasma, as can be seen from Example 10 and Comparative Example 6, the protective solution provided by the present invention can effectively improve the immunity of cells, prevent Cells infected with mycoplasma.
本发明的胎盘或脐带细胞保护液用于保存从胎盘或脐带采集后到脐带或胎盘分离前的人来源的脐带或胎盘。本发明的胎盘或脐带保护液可以在保存胎盘或脐带的过程中降低胎盘或脐带的新陈代谢，减少代谢物的累积，又能提供一些基本的营养物质和能量物质，以维持最低的代谢水平，能有效地保存胎盘或脐带中的干细胞的活性，大大减少运输、交接、检测的时间限制。经此种保护液保存的胎盘或脐带，分离出的干细胞活性受时间的影响很小，大大减少了胎盘或脐带从采集到制备的时间限制，且所用的所有成分均符合临床标准，污染率小。经此保护液2-10℃恒温保存的胎盘或脐带，在保存48小时后胎盘或脐带中细胞活性是采集的新鲜胎盘或脐带的80％，采集保存后的胎盘或脐带同不加任何溶液保存的胎盘或脐带的污染率相比，由1.2％降到了0.15％。The placenta or umbilical cord cell protection solution of the present invention is used for preserving human-derived umbilical cord or placenta from the placenta or umbilical cord collection to the umbilical cord or placenta separation. The placenta or umbilical cord protection solution of the present invention can reduce the metabolism of the placenta or umbilical cord during the preservation of the placenta or umbilical cord, reduce the accumulation of metabolites, and provide some basic nutrients and energy substances to maintain the lowest metabolic level. Effectively preserve the activity of stem cells in the placenta or umbilical cord, greatly reducing the time limit for transportation, handover, and detection. For the placenta or umbilical cord preserved in this protective solution, the activity of the isolated stem cells is little affected by time, which greatly reduces the time limit from collection to preparation of the placenta or umbilical cord, and all the ingredients used are in line with clinical standards, and the pollution rate is small . The placenta or umbilical cord stored at a constant temperature of 2-10°C in this protective solution, the cell activity in the placenta or umbilical cord after storage for 48 hours is 80% of that of the collected fresh placenta or umbilical cord, and the placenta or umbilical cord after collection and preservation are the same as those without any solution. The contamination rate of the placenta or umbilical cord dropped from 1.2% to 0.15%.
本发明不局限于上述最佳实施方式，任何人在本发明的启示下都可得出其他各种形式的产品，但不论在其形状或结构上作任何变化，凡是具有与本申请相同或相近似的技术方案，均落在本发明的保护范围之内。The present invention is not limited to the above-mentioned best implementation mode, anyone can draw other various forms of products under the inspiration of the present invention, but no matter make any changes in its shape or structure, all those with the same or similar features as the present application Approximate technical solutions all fall within the protection scope of the present invention.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107974430A (en) * | 2018-01-24 | 2018-05-01 | 北京臻溪谷医学研究中心（有限合伙） | A kind of construction method that obtains isolated culture method and stem cell bank of human adipose-derived stem cell |
| CN110903952A (en) * | 2019-11-06 | 2020-03-24 | 天晴干细胞股份有限公司 | A method for separating, purifying and resuscitating placental blood using protective fluid and placenta squeezer |
| CN112075418A (en) * | 2020-09-29 | 2020-12-15 | 广州同康生物科技有限公司 | Adipose mesenchymal stem cell cryopreservation liquid and adipose mesenchymal stem cell cryopreservation method |
| CN112970740A (en) * | 2020-12-22 | 2021-06-18 | 山东华领奥源生物科技有限公司 | Gel solution for preserving placental sub-totipotent stem cells and application thereof |
| CN113151160A (en) * | 2021-05-07 | 2021-07-23 | 天津博裕力牧科技有限公司 | Method for improving cattle in-vitro fertilization efficiency and used freezing medium and thawing medium |
| CN114190362A (en) * | 2021-12-08 | 2022-03-18 | 广东医科大学顺德妇女儿童医院(佛山市顺德区妇幼保健院) | Placenta preserving fluid and placenta preserving method |
| CN114540296A (en) * | 2022-03-22 | 2022-05-27 | 和携科技有限公司 | Preparation method of composite exosome and application of composite exosome in directionally enhancing angiogenesis capacity |
| CN116465700A (en) * | 2023-04-21 | 2023-07-21 | 上海腾瑞制药股份有限公司 | Method for detecting biological activity of active protein and polypeptide in medicine gel |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102154200A (en) * | 2010-12-13 | 2011-08-17 | 中国人民解放军第三○二医院 | Preparation and storage of mesenchymal stem cells for clinical treatment |
| CN102334472A (en) * | 2011-08-02 | 2012-02-01 | 江苏省北科生物科技有限公司 | Umbilical cord preserving fluid and preparation method thereof |
| CN103053511A (en) * | 2012-12-10 | 2013-04-24 | 山东省眼科研究所 | Cornea metaphase preservation solution, and preparing and using methods thereof |
| CN104711220A (en) * | 2015-03-13 | 2015-06-17 | 余艳春 | Novel method for preparing menstrual blood mesenchymal stem cells |
| CN104928248A (en) * | 2015-07-13 | 2015-09-23 | 北京昱龙盛世生物科技有限公司 | Kit for preparing neural stem cells and method for preparing neural stem cells |
| CN105076114A (en) * | 2015-08-21 | 2015-11-25 | 深圳爱生再生医学科技有限公司 | Serum-free preservation liquid for umbilical cord mesenchymal stem cells and application of serum-free preservation liquid |
| CN205062086U (en) * | 2015-09-18 | 2016-03-02 | 北京昱龙盛世生物科技有限公司 | Cultivate kit of nerve cord cell |
| CN105586309A (en) * | 2016-02-06 | 2016-05-18 | 福建省银丰干细胞工程有限公司 | Method for acquiring safe and effective umbilical cord mesenchymal stem cell |
-
2016
- 2016-08-01 CN CN201610625635.XA patent/CN107114356A/en active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102154200A (en) * | 2010-12-13 | 2011-08-17 | 中国人民解放军第三○二医院 | Preparation and storage of mesenchymal stem cells for clinical treatment |
| CN102334472A (en) * | 2011-08-02 | 2012-02-01 | 江苏省北科生物科技有限公司 | Umbilical cord preserving fluid and preparation method thereof |
| CN103053511A (en) * | 2012-12-10 | 2013-04-24 | 山东省眼科研究所 | Cornea metaphase preservation solution, and preparing and using methods thereof |
| CN104711220A (en) * | 2015-03-13 | 2015-06-17 | 余艳春 | Novel method for preparing menstrual blood mesenchymal stem cells |
| CN104928248A (en) * | 2015-07-13 | 2015-09-23 | 北京昱龙盛世生物科技有限公司 | Kit for preparing neural stem cells and method for preparing neural stem cells |
| CN105076114A (en) * | 2015-08-21 | 2015-11-25 | 深圳爱生再生医学科技有限公司 | Serum-free preservation liquid for umbilical cord mesenchymal stem cells and application of serum-free preservation liquid |
| CN205062086U (en) * | 2015-09-18 | 2016-03-02 | 北京昱龙盛世生物科技有限公司 | Cultivate kit of nerve cord cell |
| CN105586309A (en) * | 2016-02-06 | 2016-05-18 | 福建省银丰干细胞工程有限公司 | Method for acquiring safe and effective umbilical cord mesenchymal stem cell |
Non-Patent Citations (5)
| Title |
|---|
| 徐文婧: "影响脐带间充质干细胞活性的基质及其保护液的研究", 《中国优秀硕士学位论文全文数据库 医药卫生科技辑》 * |
| 胡春: "《低温医学》", 30 September 1993, 人民卫生出版社 * |
| 郭立达等: "《动物细胞分离培养》", 31 August 2015, 重庆大学出版社 * |
| 鄂征: "《组织培养和分子细胞学技术》", 31 December 1994, 北京出版社 * |
| 顾其胜等: "《实用生物医用材料学》", 30 September 2005, 上海科学技术出版社 * |
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| CN110903952A (en) * | 2019-11-06 | 2020-03-24 | 天晴干细胞股份有限公司 | A method for separating, purifying and resuscitating placental blood using protective fluid and placenta squeezer |
| CN112075418A (en) * | 2020-09-29 | 2020-12-15 | 广州同康生物科技有限公司 | Adipose mesenchymal stem cell cryopreservation liquid and adipose mesenchymal stem cell cryopreservation method |
| CN112075418B (en) * | 2020-09-29 | 2021-10-22 | 惟力维斯(上海)医学科技有限公司 | Adipose-derived mesenchymal stem cell cryopreservation solution and method for cryopreservation of adipose-derived mesenchymal stem cells |
| CN112970740A (en) * | 2020-12-22 | 2021-06-18 | 山东华领奥源生物科技有限公司 | Gel solution for preserving placental sub-totipotent stem cells and application thereof |
| CN113151160A (en) * | 2021-05-07 | 2021-07-23 | 天津博裕力牧科技有限公司 | Method for improving cattle in-vitro fertilization efficiency and used freezing medium and thawing medium |
| CN113151160B (en) * | 2021-05-07 | 2022-05-13 | 天津力牧生物科技有限公司 | Method for improving the efficiency of bovine in vitro fertilization and cryopreservation and thawing solutions used |
| CN114190362A (en) * | 2021-12-08 | 2022-03-18 | 广东医科大学顺德妇女儿童医院(佛山市顺德区妇幼保健院) | Placenta preserving fluid and placenta preserving method |
| CN114540296A (en) * | 2022-03-22 | 2022-05-27 | 和携科技有限公司 | Preparation method of composite exosome and application of composite exosome in directionally enhancing angiogenesis capacity |
| CN114540296B (en) * | 2022-03-22 | 2023-11-24 | 和携科技有限公司 | Preparation method of composite exosome and application of composite exosome in directional enhancement of angiogenesis capacity |
| CN116465700A (en) * | 2023-04-21 | 2023-07-21 | 上海腾瑞制药股份有限公司 | Method for detecting biological activity of active protein and polypeptide in medicine gel |
| CN116465700B (en) * | 2023-04-21 | 2023-11-28 | 上海腾瑞制药股份有限公司 | Method for detecting biological activity of active protein and polypeptide in medicine gel |
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