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| 产品名称 | UltraBio™大分子量蛋白预制胶（Tris-Acetate） | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| 产品介绍 |
阿拉丁的UltraBio™大分子量蛋白预制胶(Tris-Acetate) (UltraBio™ Tris-Acetate Precast PAGE Gel for High Molecular Weight Protein)是一种主要用于大分子量蛋白(40-500kDa)检测的使用安全、便捷、高品质的常规尺寸聚丙烯酰胺预制凝胶，采用Tris-Acetate SDS (SDS-PAGE)或Tris-Glycine (Native-PAGE)电泳缓冲系统可以分别进行变性和非变性蛋白凝胶电泳，仅需约60分钟即可完成电泳并获得非常平整锐利的条带。本预制胶有1.5厘米高的浓缩胶，具有非常优良的分离效果，电泳后蛋白条带平整、清晰、细腻、锐利，几乎没有边缘效应；同时本预制胶胶板为玻璃材质，减少了对蛋白的非特异性吸附，电泳效果非常好，达到甚至超过了自配PAGE胶的电泳效果；兼容市场上主流的小型电泳槽，常用于PAGE和Western检测。 阿拉丁的UltraBio™大分子量蛋白预制胶(Tris-Acetate)提供3-8%浓度的梯度胶和7%浓度的固定浓度胶，并有10孔和15孔两种孔数选择。每种预制胶的最佳分离范围请参考下表：
本预制胶含有1.5厘米高的4%浓缩胶，可以有效确保获得非常锐利的条带。 本预制胶丙烯酰胺(Acrylamide)与甲叉丙烯酰胺(Bisacrylamide)的比例为29:1，凝胶厚度为1.5mm。加样孔数为10孔的最大上样量为60μl，加样孔数为15孔的最大上样量为30μl。胶板尺寸：宽×高×厚度为98×84×4.1mm；凝胶尺寸：宽×高×厚度为81×74×1.5mm。 聚丙烯酰胺凝胶电泳(Polyacrylamide gel electrophoresis, PAGE)技术广泛用于蛋白质、核酸及蛋白质-核酸复合物的分离纯化、检测、鉴定、分子量分析等实验，是生命科学研究中最基本的实验技术之一。常见的Western印迹(Western blot)检测就是基于PAGE的。Glycine-SDS-PAGE (也被称为Laemmli-SDS-PAGE，基于Tris-Glycine缓冲系统)和Tricine-SDS-PAGE (基于Tris-Tricine缓冲系统)是广泛应用的将蛋白质按照分子量大小分离的方法。Tricine-SDS-PAGE可以很好地分离小于30kDa的蛋白质，而Glycine-SDS-PAGE被建议用于大于30kDa的蛋白。不同体系分离蛋白质的特性与Glycine和Tricine的pKa值有关，不同大小的蛋白质在不同的缓冲体系中拥有不同的电泳迁移率，且特定范围内的蛋白质分辨率较高。因此，如果需要分离大分子量蛋白，就需要选择合适的电泳系统。同时，也建议使用由下至上丙烯酰胺浓度降低的梯度胶。目前研究中常用Tris-Acetate凝胶系统分离大分子量蛋白。在Tris-Acetate凝胶系统中，Acetate(-)来自凝胶缓冲液，相较于系统中其它的阴离子，Acetate(-)对阳极具有极高的亲和力，因此可作为前导离子(Leading ion)。凝胶缓冲液中含有Tris(+)和Acetate(-)，pH为7.0，使凝胶具有很高的分辨率。电泳缓冲液中含Tris(+)、Tricine(-)和十二烷基硫酸盐(Dodecylsulfate, DS)，其中Tricine(-)作为尾随离子(Tailing ion)。电泳缓冲液pH为8.24，减少了蛋白修饰，使条带更锐利。对高分子量蛋白进行分离的结果也表明，Tris-Acetate凝胶分离效果好、转移效率高、灵敏度高、分辨率佳。此外，Tris-Acetate凝胶也被报道可以作为一种稳定的、经济高效的研究蛋白质寡聚化的工具，仅使用单一凝胶即可提供出色的蛋白质寡聚化结果。 阿拉丁的UltraBio™大分子量蛋白预制胶(Tris-Acetate)使用中性pH的Tris-Acetate缓冲液制备，不含SDS，既可用于变性蛋白电泳，也可用于非变性蛋白电泳。 本预制胶电泳后可以使用Tris-Glycine缓冲系统的转膜液进行转膜，但需降低转膜液中乙醇或甲醇浓度(至5%)。 关于10孔和15孔预制胶的选择：需要检测的样品数量多或者需要定量时，推荐使用15孔预制胶，通量更大、更便于进行较多样品的定量统计分析；需要获得非常漂亮的代表性图片时，推荐使用10孔预制胶，10孔预制胶获得的条带更加平整和锐利。 本产品使用安全、便捷。本预制胶无需配制，即开即用，去掉梳子即可上样，而传统的PAGE配制凝胶繁琐费时，并且制胶时还会接触有毒和刺激性试剂。 本产品质量稳定。本预制胶采用高品质玻璃胶板，和塑料胶板相比，大大减少了胶板对蛋白的吸附，电泳效果更好。本产品流水线灌注，品质稳定可靠，重复性好，不同批次的产品一致性高。 本产品电泳效果好。本预制胶的蛋白质分离效果极佳，蛋白条带平整、清晰、细腻、锐利，转膜效率高。 本产品电泳时间短。本预制胶推荐的变性电泳电压和电泳时间为150V 50-70分钟；非变性电泳电压和电泳时间为150V 90-180分钟。完成电泳后可获得非常平整和锐利的电泳条带。具体的电泳时间可以根据溴酚蓝的电泳位置或实验的具体需求进行确定。 本产品取出凝胶极为便捷。只需用刀片在玻璃胶板一侧轻轻划一下即可，并且玻璃胶板打开极为方便，无需特殊的起撬工具。
注意事项： 本预制胶不能置于0℃以下冷冻，否则凝胶会冻裂。 内槽电泳液和转膜液建议新鲜配制，试剂纯度不够、反复使用或长期放置的缓冲液会降低电泳效果。 电泳结束后可使用Tris-Glycine缓冲系统的转膜液进行转膜。乙醇或甲醇会使大分子蛋白很容易沉淀。可通过减少转膜液中的乙醇或甲醇百分比(至5%)来避免这种情况发生。为了进一步确保蛋白质不会沉淀，可添加SDS至终浓度为0.1%。SDS向蛋白添加均匀的负电荷，使得它们更容易从凝胶转移到膜上。 转膜时，推荐使用0.45μm的PVDF膜。 本产品仅限于专业人员的科学研究用，不得用于临床诊断或治疗，不得用于食品或药品，不得存放于普通住宅内。 为了您的安全和健康，请穿实验服并戴一次性手套操作。 The UltraBio™ Tris-Acetate Precast PAGE Gel for High Molecular Weight Protein is a safe, high-quality, and ready-to-use polyacrylamide gel primarily used to analyze high molecular weight proteins (40-500kDa). It can be used for both denaturing and native protein electrophoresis with the Tris-Acetate SDS (SDS-PAGE) and Tris-Glycine (Native-PAGE) running buffers, respectively. It takes approximately 60 minutes to complete electrophoresis, and well-separated and sharp protein bands can be obtained. This precast gel has a 1.5cm high 4% stacking gel, with a very good protein separation effect. Moreover, this product uses glass plates to reduce non-specific adsorptions. This product has identical or even better performance than home-made PAGE gels and can be routinely used for PAGE and Western analysis. Aladdin provides both UltraBio™ precast gradient/fixed gels of different concentrations with 10 or 15 loading wells. The gradient gel concentration is 3-8%, and the fixed gel concentration is 7%. Please refer to the table below for the optimal protein separation range for the UltraBio™ Tris-Acetate Precast PAGE Gels.
This product has a 4% stacking gel with a height of 1.5cm to ensure very sharp protein bands can be obtained. The ratio of acrylamide to bisacrylamide is 29:1, and the gel thickness is 1.5mm. The maximum loading volume per well of 10-well and 15-well gels is 60μl and 30μl, respectively. The plate size is 98×84×4.1mm (width × height × thickness), with a gel size of 81×74×1.5mm (width × height × thickness). Polyacrylamide gel electrophoresis (PAGE) is a widely used technique in life science research for the separation, purification, assay, identification, and molecular weight analysis of proteins, nucleic acids, and protein-nucleic acid complexes. The commonly used Western blot technique is based on PAGE. Glycine-SDS-PAGE (also known as Laemmli-SDS-PAGE, based on the Tris-Glycine buffer system) and Tricine-SDS-PAGE (based on the Tris-Tricine buffer system) are widely used methods for separating proteins based on their molecular weight. Tricine-SDS-PAGE is particularly effective in separating proteins smaller than 30kDa, while Glycine-SDS-PAGE is recommended for proteins larger than 30kDa. The separation characteristics of proteins in different systems are related to the pKa values of Glycine and Tricine. Proteins of different sizes have different electrophoretic mobilities in different buffer systems, and proteins within specific size ranges can be resolved with high resolution. Therefore, when separating high molecular weight proteins, it is important to choose the appropriate electrophoresis system. It is also recommended to use gradient gels with decreasing acrylamide concentration from the bottom to the top. Currently, the Tris-Acetate gel system is commonly used for separating high molecular weight proteins. In the Tris-Acetate gel system, Acetate (-) comes from the gel buffer, and compared to other anions in the system, Acetate (-) has a high affinity for the anode and can serve as the leading ion. The gel buffer contains Tris (+) and Acetate (-) at a pH of 7.0, which provides high resolution in the gel. The electrophoresis buffer contains Tris (+), Tricine (-), and dodecyl sulfate (DS), with Tricine (-) acting as the trailing ion. The pH of the electrophoresis buffer is 8.24, which reduces protein modifications and results in sharper bands. The results of separation of high molecular weight proteins have also shown that Tris-Acetate gels provide good separation, high transfer efficiency, high sensitivity and excellent resolution. In addition, Tris-Acetate gels have been reported to serve as a stable and cost-effective tool for the study of protein oligomerization, providing excellent protein oligomerization results using only a single gel. Unlike traditional Tris-Glycine gels, UltraBio™ Tris-Acetate Precast PAGE Gels are prepared with neutral pH tris-acetate buffer and do not contain SDS. They can be used for both denaturing and native protein electrophoresis.
After electrophoresis, protein can be transferred from the gel to blot membrane with the Tris-Glycine buffer system.However, the concentration of ethanol or methanol in the transfer buffer needs to be reduced to 5%.
Selection of 10-well and 15-well precast gels: Protein bands obtained by 10-well precast gels are flatter and sharper. When there is a large number of samples to be examined or protein quantification is required, we recommend using the 15-well precast gel which offers greater throughput and is more convenient for statistical quantitation analysis. This product is safe and convenient to use. There is no need for gel preparation and casting, thus avoiding contact with toxic and irritating reagents. After electrophoresis, the gel can be easily removed from the glass plates. The quality of this product is stable, with high consistency between batches. This product has good protein resolution performance. Proteins in the gel can be transferred efficiently as well. The electrophoresis time is short. The denaturing electrophoresis can be completed in 50-70 minutes at 150V, while the native electrophoresis can be completed in 90-180 minutes at 150V. The run time varies depending on experimental requirements.
Precautions: Do NOT freeze this product at temperatures below 0℃. The electrophoresis buffer in the inner chamber of the electrophoresis tank and the transfer buffer should be freshly prepared. Low-quality buffers, recycled buffers, or buffers that have been kept for too long period of time will result in poor protein resolution and transfer. After electrophoresis, protein transfer can be performed using the Tris-Glycine transfer buffer system. Ethanol or methanol can cause precipitation of large protein molecules, which can be avoided by reducing the percentage of ethanol or methanol in the transfer buffer to 5%. SDS can be additionally added to a final concentration of 0.1% to further ensure no precipitation of proteins. SDS enables a uniform negative charge of proteins, allowing them to be more easily transferred from the gel to the membrane. For transfer, we recommend using 0.45μm PVDF Membrane. This product is for R&D only. Not for drug, household, or other uses. For your safety and health, please wear a lab coat and disposable gloves during the operation. |
储存与运输
| 储存温度 | 2-8°C储存 |
|---|---|
| 运输条件 | 冰袋运输 |
| 稳定性与储存 | 4℃保存，1-2个月有效。切勿置于0℃以下冷冻。 |
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