H2303384计算溶液所需的质量、体积或浓度。
为什么选择此级别
，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
避光,-20°C储存。超低温运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 1 篇同行评审文献引用。
概述
荧光染料PKH67 适用于常规细胞膜标记，是⼀种可对体外和体内细胞示踪的绿色荧光染料，通过与膜结构的脂质分⼦结合⽽标记细胞。PKH67对细胞毒性较⼩，荧光背景低，脂溶性⾼，能够轻易穿透细胞膜，有着强⽽稳定的绿⾊荧光。经PKH67标记的细胞可⽤于体外和体内增殖研究，且具有不会使邻近细胞染⾊的功能。在细胞分裂增殖过程中，PKH67 的荧光强度会随着细胞的分裂⽽逐级递减，标记荧光可平均分配⾄两个⼦代细胞中，因此其荧光强度是亲代细胞的⼀半，根据这⼀特性，它可被⽤于检测细胞增殖，细胞周期的估算及细胞分裂等⽅⾯。PKH67标记细胞的荧光⾮常均⼀，并且分裂后的⼦代细胞的荧光分配也更均⼀。在细胞分裂增殖过程中，PKH67标记荧光可平均分配⾄两个⼦代细胞中，荧光强度变为亲代细胞的⼀半，通过流式细胞仪根据荧光强度的不同，可检测出未分裂细胞，分裂⼀次(1/2 的荧光强度)，⼆次(1/4 的荧光强度)，三次(1/8的荧光强度)，以及更多分裂次数的细胞。PKH67 可检测分裂次数多达六次甚⾄更多。除了⽤于细胞增殖检测，PKH67 还可以⽤于细胞的体外盒体内⽰踪，标记后荧光在胞内表达稳定，阳性标记率达98%以上，标记细胞形态良好，能有效地观察细胞在体外的诱导分化情况；或将标记的细胞注⼊体内，可以有效的显⽰移植细胞在活体组织中的迁移及分化。PKH67标记的细胞⽤于体内观察可以长达数周之久，它常被⽤来做活体细胞检测实验和⽤荧光电镜观察细胞长期活动的实验。PKH67 毒性较⼩，不影响细胞的增殖能⼒。此⽅法操作简单，且不⽤放射性同位素，不存在安全隐患。可以更快速，更准确和更安全地得到想要的实验数据。
由于炭尾长度更长，内部研究已经证明PKH67比PKH2由更少的细胞间转移。在采用PKH1和PKH2进行的体内研究中，荧光强度都会缓慢损失。由于这是绿色细胞linker染料而非红色细胞linker染料出现的行为特征，因而PKH67会出现类似的性质。不分裂细胞的体外细胞膜留存和体内荧光半衰期的关联性揭示，PKH67的体内荧光半衰期为10-12天。其他具有类似半衰期的绿色细胞linker染料已经被用于监测1-2月内的体内 淋巴细胞和巨噬细胞运输，结果表明PKH67还可用于中等时长的体内跟踪研究。
染料可以稳定的与细胞膜脂质区结合并发出荧光，主要用于细胞体外标记、体外细胞增殖研究以及体内外的细胞示踪研究。PKH67的体内荧光半衰期为10-12天。相比于PKH-67，PKH-26具有更长的半衰期，标记在兔红细胞上的PKH26半衰期长达100天以上。特别适用于体外增殖研究以及长期的体内细胞跟踪研究。PKH67标记细胞后通常⽤流式细胞仪进⾏细胞增殖检测。
试剂盒组分
0.1ml套装：P266290A-0.1ml P266290B-10ml
1ml套装： P266290A-1ml P266290B-60ml
带A后缀的是染料，B后缀的是稀释液，染料和稀释液配套使用
PKH67标记细胞呈绿⾊荧光，荧光波长：λex=490 nm, λem=502 nm。
储存条件：-20℃避光保存有效期1年
注意事项
●染⾊浓度根据细胞种类的不同和每孔内细胞数量的多少⽽异。
● 配制的PKH67母液极易⽔解，建议分装保存， ≦-20℃冷冻干燥保存。
● PKH67 ⼯作液应现配现⽤，不能提前配制，因为PKH67 吸⽔会分解，影响染⾊效果。
● PKH67 易被⽔解，在⽔溶液中会很快变质。母液请在使⽤过程中避免接触⽔。⼯作液在标记细胞的过程中和⽔接触是在许可的时间范围内的。
● PKH67荧光染⾊剂为DMSO溶液，在4℃、冰浴等较低温度情况下会凝固⽽粘在离⼼管管底、管壁或管盖内，从冰箱取出后恢复⾄室温，变成液体状态后离⼼⾄管底部再开盖。可以37℃⽔浴⽚刻⾄全部融解后使⽤。
● 不同的细胞种类标记后可以⽰踪的代次或时间差异较⼤，请根据实际情况或参考⽂献进⾏检测。
使用方法
1．染色液制备：
（1）从冰箱中取出PKH67试剂，静置⼏分钟⾄室温，或者37℃⽔浴⽚刻后，离⼼盛放PKH67 的管⼦，开盖前请务必离⼼⼏分钟让试剂充分落⼊管底后才能开盖。
（2）根据需要检测的细胞样品数，⽤稀释液将探针10倍稀释，再⽤合适的溶液(如：⽆⾎清培养基，HBSS或PBS)将PKH67母液25倍稀释，配制成染⾊⼯作液。最佳⼯作液浓度请根据不同细胞和⾃⾝的实验体系来调整。⼀般细胞使⽤按试剂盒中的母液的终浓度250倍稀释即可，有些细胞可能需要适当增加浓度。
2. 细胞染色
（1）将制备好的待测细胞⽤100μl染⾊⼯作液重悬，⾄细胞浓度⼤约107/ml。也可以进⾏原位染⾊，染⾊液⾜够覆盖细胞即可。
（2）在2～8 ℃培养细胞 15～30分钟，不同的细胞最佳培养时间不同。
建议待标记细胞在染⾊⼯作液中于37℃孵育5min，再于4℃孵育15min。
低温孵育可降低细胞对染料的内吞作⽤，有助于染料对质膜的标记，并且降低染料定位细胞质囊泡的可能性。
（3）离⼼后去上清，收集细胞，⽤PBS或⽆⾎清培养基洗涤细胞1-2次，最后加⼊PBS或⽆⾎清培养基重悬细胞。
（4）取500μl 细胞悬液，⽤流式细胞仪检测。Ex/Em=490/502nm。
（5）随后还可按照细胞的正常培养⽅法进⾏培养。
（6）可以在荧光显微镜下直接观察标记效果，也可以在培养适当时间后再⽤流式细胞仪检测细胞增殖，或⽤于其他特定实验⽬的的细胞荧光⽰踪。
The fluorescent dye PKH67 is suitable for conventional cell membrane labeling. It is a green fluorescent dye that can track cells in vitro and in vivo. It labels cells by binding to the lipid components of the membrane structure. PKH67 has low cytotoxicity, low fluorescence background, high fat solubility, can easily penetrate cell membranes, and has strong and stable green fluorescence. PKH67-labeled cells can be used for in vitro and in vivo proliferation studies, and have the function of not staining neighboring cells. In the process of cell division and proliferation, the fluorescence intensity of PKH67 will gradually decrease as the cells divide. The labeled fluorescence can be evenly distributed to the two sub-generation cells, so its fluorescence intensity is half that of the parent cell. According to this feature , It can be used to detect cell proliferation, cell cycle estimation and cell division, etc. The fluorescence of PKH67-labeled cells is very uniform, and the fluorescence distribution of sub-generation cells after division is also more uniform. In the process of cell division and proliferation, PKH67-labeled fluorescence can be evenly distributed between the two sub-generation cells, and the fluorescence intensity becomes half of that of the parent cell. According to the difference in fluorescence intensity, the undivided cells can be detected by flow cytometry. One time (1/2 the fluorescence intensity), the second time (1/4 the fluorescence intensity), three times (1/8 the fluorescence intensity), and more divisions of cells. PKH67 can detect splits up to six times or even more. In addition to the detection of cell proliferation, PKH67 can also be used for in vitro tracking of cells. After labeling, the fluorescence expression is stable in the cell, and the positive labeling rate is over 98%. The labeled cells have good morphology, which can effectively observe the cells in vitro. Induce differentiation; or inject labeled cells into the body, it can effectively show the migration and differentiation of transplanted cells in living tissues. PKH67-labeled cells can be used for in vivo observation for as long as several weeks. It is often used for in vivo cell detection experiments and experiments to observe long-term cell activity using fluorescence electron microscope. PKH67 is less toxic and does not affect cell proliferation. This method is simple to operate, does not use radioactive isotopes, and poses no safety hazards. You can get the desired experimental data faster, more accurately and more safely.
Due to the longer length of the charcoal tail, internal studies have shown that PKH67 is less transferred between cells than PKH2. In in vivo studies using PKH1 and PKH2, the fluorescence intensity will slowly lose. Since this is a behavioral characteristic of green cell linker dye rather than red cell linker dye, PKH67 will have similar properties. The correlation between the in vitro cell membrane retention of non-dividing cells and the in vivo fluorescence half-life reveals that the in vivo fluorescence half-life of PKH67 is 10-12 days. Other green cell linker dyes with similar half-lives have been used to monitor the transport of lymphocytes and macrophages in the body within one to two months. The results indicate that PKH67 can also be used for medium-term in vivo tracking studies.
The dye can stably bind to the lipid region of the cell membrane and emit fluorescence, and is mainly used for cell labeling in vitro, cell proliferation research in vitro, and cell tracing research in vivo and in vitro. The fluorescence half-life of PKH67 in vivo is 10-12 days. Compared with PKH-67, PKH-26 has a longer half-life, and the half-life of PKH26 labeled on rabbit red blood cells is more than 100 days. Especially suitable for in vitro proliferation research and long-term in vivo cell tracking research. After PKH67 labels the cells, flow cytometry is usually used for cell proliferation detection.
Kit components
0.1ml kits: P266290A-0.1ml P266290B-10ml
1ml kits: P266290A-1ml P266290B-60ml
Dyes with A suffix and diluents with B suffix are used together
PKH67 labeled cells show green fluorescence, the fluorescence wavelength: λex=490 nm, λem=502 nm.
Storage conditions: -20℃ protected from light, valid for 1 year
Precautions
●Staining concentration varies according to the type of cell and the number of cells in each well.
● The prepared PKH67 mother liquor is very easy to dissolve. It is recommended to store in aliquots and freeze-dry at ≦-20℃.
● PKH67 working solution should be prepared for immediate use, and cannot be prepared in advance, because PKH67 will decompose due to the absorption of water and affect the dyeing effect.
● PKH67 is easily decomposed and will deteriorate quickly in the water solution. Please avoid contact with water during use of mother liquor. The working fluid is in contact with the water during the process of labeling the cells within the permitted time range.
● PKH67 fluorescent dye is a DMSO solution. It will solidify and stick to the bottom, wall or cap of the tube at a lower temperature such as 4℃ and ice bath. After being taken out of the refrigerator, it will return to room temperature and become After the liquid is in the state, remove the cap from the bottom of the tube. It can be used after it has completely melted in a 37°C water bath.
● The number of generations or time that can be traced after different cell types are marked is quite different. Please make a test based on the actual situation or reference documents.
Instructions
1. Staining solution preparation:
(1) Take out the PKH67 reagent from the refrigerator, let it stand for a few minutes to room temperature, or after a 37°C water bath, leave the tube containing PKH67, and be sure to leave the tube for a few minutes before opening the lid to allow the reagent to fully fall into the tube The lid can only be opened after the bottom.
(2) According to the number of cell samples to be tested, dilute the probe 10 times with the diluent, and then use a suitable solution (such as non-clear medium, HBSS or PBS) to dilute the PKH67 mother liquor 25 times to prepare a stain Work fluid. The best working solution concentration should be adjusted according to different cells and your own experimental system. Generally, the cells can be diluted 250 times according to the final concentration of the mother liquor in the kit. Some cells may need to increase the concentration appropriately.
2. Cell staining
(1) Resuspend the prepared cells to be tested in 100μl of staining solution to a cell concentration of about 107/ml. You can also perform in-situ staining, as long as the staining solution is enough to cover the cells.
(2) Culture the cells at 2～8℃ for 15～30 minutes. The best culture time is different for different cells.
It is recommended to incubate the labeled cells in the staining solution at 37°C for 5 minutes, and then at 4°C for 15 minutes.
Low-temperature incubation can reduce the endocytosis of the dye by the cells, help the dye to label the plasma membrane, and reduce the possibility of the dye localizing to cytoplasmic vesicles.
(3) After separation, remove the supernatant, collect the cells, wash the cells 1-2 times with PBS or non-clear medium, and finally add PBS or non-clear medium to resuspend the cells.
(4) Take 500μl of cell suspension and test with flow cytometer. Ex/Em=490/502nm.
(5) Subsequently, the cells can be cultured according to the normal culture method.
(6) The labeling effect can be directly observed under a fluorescence microscope, or the cell proliferation can be detected by a flow cytometer after an appropriate period of culture, or used for cell fluorescence traces for other specific experimental purposes.
规格
英文名称
PKH67 Green Fluorescent Cell Linker Mini Kit
储存与运输
储存条件
避光,-20°C储存
运输条件
超低温运输
技术文档
高级数据
质检证书（CoA,COO,BSE/TSE 和分析图谱)
此产品的引用文献
溶液计算器
产品仅提供给经过验证的企业、机构和合格的专业人员用于研发。不适用于人类、动物、诊断或治疗。
Copyright © 2023至今 上海阿拉丁生化科技股份有限公司 保留所有权利。
危险品化学品经营许可证（带存储）