UltraBio™ Transfer Buffer

UltraBio™ 转移缓冲液

T743350-1L

1L

Suitable for Bis-Tris Gels

Powder, for Bis-Tris Gels

阿拉丁生产的UltraBio™ Transfer Buffer (Powder, for Bis-Tris Gels)，即UltraBio™ Bis-Tris胶专用转膜液(Powder)，是一种粉末形态的安全用于Western时湿法转膜的缓冲液。转膜液也称转印缓冲液或电转液，本转膜液专用于Bis-Tris胶，也可用于Tris-Acetate胶。本转膜液可在转膜过程中维持中性pH环境，从而保护氨基酸侧链不受改性，防止氨基酸侧链修饰，与使用Edman化学降解法的蛋白质N-端测序技术兼容1]。对还原性蛋白进行转膜时，可在本转膜液中加入0.1%的阿拉丁P0737 UltraBio™ Antioxidant (400X)，以促进还原性蛋白向膜转移。本转膜液由高质量的Bicine、Bis-Tris、EDTA等试剂混合而成，为粉末装，配制时无需调整pH值。如需20倍浓缩装，推荐使用阿拉丁UltraBio™ Transfer Buffer (20X, for Bis-Tris Gels) 。对于Bis-Tris胶和Tris-Acetate胶，也可以使用Tris-Glycine缓冲系统的Western转膜液进行转膜。本转膜液可以回收，回收后可以再使用1-2次。1瓶本产品可配制1L转膜液。室温保存，一年有效。不推荐将还原性蛋白样品和非还原性蛋白样品一起进行转膜。对还原性蛋白进行转膜时，可在配制转膜液时加入0.1%的UltraBio™ Antioxidant (400X) 。含有10%乙醇或甲醇的转膜液可在印迹模块中实现单块凝胶的最佳转移效果。如果需转移2块凝胶，请将乙醇或甲醇含量增加到20%，以确保蛋白的有效转移。本产品适用于Bis-Tris和Tris-Acetate凝胶系统的转膜液，不适用于其它凝胶系统。如果转膜时设定的电流或电压较大，会有比较明显的产热现象，即产生所谓的焦耳热效应(Joule heating)，并导致转膜液的电阻变小，最终影响转膜效率、转膜液的缓冲能力、甚至过热时导致凝胶熔化或黏附在膜上。因此建议转膜时，在转膜槽内用冰盒降温，并将转膜槽置于冰浴中。同时也可以考虑转膜液配制好后先在4℃或者冰浴预冷。使用本产品转膜时，如果设定恒流为300mA，无论是同一个转膜槽中1或2块胶转膜，电压在开始时一般为100V左右。由于转膜过程中，转膜液温度会逐渐升高，电阻变小，所以结束时电压一般为120V左右。转膜效率和电场强度(Electric field strength, 用E表示)相关，而电场强度主要和电压及电极之间的距离相关，即E=U/d。由于用于转膜的电源品牌和型号的不同，一方面最高电流差异较大，另一方面功率也会有较大差别。通常大功率的电源相对贵一些，也更适合用于转膜。比如Bio-Rad Powerpac Basic Power Supply的最大功率是75W，在设定恒流为400mA时，电压超过187V时就会超过电源的最大功率而出现过载保护。有些电源会自动降低电压或电流，有些则会报错并停止转膜。对于出现停止转膜的情况，需要调低电流或电压，或者选择功率更大的电源。转膜时转膜液的温度、凝胶的参数(如数量、厚度、离子浓度、丙烯酰胺浓度等)、滤纸的厚度、同时使用的转膜槽数量、转膜液的离子浓度、转膜液的体积、转膜液使用的次数等都可能会影响电流和电压。其中一个电泳同时使用的转膜槽数量是最关键因素，如果是恒流，每增加一个转膜槽，每个槽的电流和电压都会相应减少一份。如果需要同时使用2个或以上的转膜槽，建议使用高电流的大功率电源，或者设定较低的电流或电压进行比较长时间的转膜。转膜时，如果出现报错而停止，需要根据情况查看是否是电压或电流过载，或者是如转膜液过多或过少、转膜槽的短路或断路等异常情况。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。 使用说明 ： 1.请按照下表配制转膜液。 Reagent Reduced Samples Non-Reduced Samples Transfer Buffer (Powder, for Bis-Tris Gels) 1 bottle 1 bottle Antioxidant (400X) 1ml - EthanolorMethanol 100-200ml 100-200ml H2O To1000ml To1000ml Total Volume 1000ml 1000ml 注：如果配制好的转膜液使用前能4℃预冷，转膜效果更好。没有用完的转膜液可在常温或4℃保存，通常两周内可以正常使用。当转膜液颜色变为浅棕色或黄褐色时，应该丢弃。2.转膜条件：300mA恒流或100V恒压1小时(推荐使用外置冰浴)即可很好地完成转膜。对于小分子量蛋白，可以适当减少转膜时间；同样对于大分子量蛋白，可以适当增加转膜时间。本转膜液也适合于快速转膜，如500mA恒流15分钟即可。3.详细的Western操作可以参考阿拉丁相关网页：http://www.aladdin-e.com/support/western.htm。参考文献：1.Chang E, Pourmal S, Zhou C, Kumar R, Teplova M, Pavletich NP, Marians KJ, Erdjument-Bromage H. J Biomol Tech. 2016. 27(2):61-74. Aladdin's UltraBio™ Transfer Buffer (Powder, for Bis-Tris Gels), also known as UltraBio™ Bis-Tris Adhesive Transfer Liquid (Powder), can be used for wet transfer during Western blot. This product is specially designed for use with Bis-Tris gels, but can also be used for Tris-Acetate gels.This product can maintain a neutral pH environment during the transfer, thereby protecting the amino acid side chain from modification. It is compatible with the Edman degradation technology to determine the N-terminal amino acid sequences of proteins [1].When transferring reducing proteins, it is recommended to add 0.1% of Aladdin's UltraBio™ Antioxidant (400X) in this transfer buffer to promote the transfer of reducing proteins.This transfer buffer prepared with high-quality Bicine, Bis-Tris, EDTA and other reagents, and there is no need to adjust the pH value for preparation. Investigators can also purchase the UltraBio™ Transfer Buffer (20X, for Bis-Tris Gels) for more convenient use.For protein transfer from Bis-Tris and Tris-Acetate gels, it is also feasible to use the Western Transfer Buffer with the Tris-Glycine buffer system.The transfer buffer prepared with this product can be recycled for 1-2 times.One bottle of this product is sufficient for preparing of 1L of transfer buffer. Precautions ： We do not recommend transferring reducing protein samples and non-reducing protein samples together. For transferring reducing proteins, it is recommended to add 0.1% Antioxidant (400X) (, P0737) in the transfer buffer.Transfer buffer containing 10% ethanol or methanol enables optimal protein transfer of a single gel. For simultaneous protein transfer from two gels, increase the ethanol or methanol content to 20% to ensure efficient transfer of proteins.This product is suitable for Bis-Tris and Tris-Acetate gel systems, not suitable for gels with other buffer systems.If the current or voltage during the transfer is high, the generated Joule heating leads to a lower resistance of the transfer buffer, which ultimately affects the transfer efficiency, the buffering capacity of the transfer buffer, and even causes the gel to melt or adhere to the membrane. Therefore, we recommend performing the transfer in an ice box or at 4℃.When using this product for protein transfer and setting the constant current at 400mA, the voltage is generally 150-200V at the beginning no matter there is 1 or 2 pieces of membrane in the transfer tank. As the temperature of the transfer buffer will gradually increase and the resistance will become smaller, the voltage is generally 100-150V at the end. The transfer efficiency is related to the electric field strength (E) which is mainly related to the voltage and the distance between the electrodes, i.e., E=U/d.If an error occurs during the transfer, check whether it is an overload of voltage or current, or abnormal conditions such as too much or too little transfer buffer, short circuit or breakage of the transfer tank, etc. Some power supplies will automatically reduce the voltage or current, while others will report an error and stop running. When the transfer stops, it is necessary to reduce the current or voltage, or choose a power supply with more power.The temperature of the transfer buffer during the transfer, the parameters of the gel (e.g., number, thickness, ionic concentration, acrylamide concentration, etc.), the thickness of the filter paper, the number of transfer tanks used simultaneously, the ionic concentration of the transfer buffer, the volume of the transfer buffer, and the number of times the transfer buffer is used may all affect the current and voltage. Among them, the number of simultaneous transfer tanks used for one electrophoresis is the most critical factor. In case of constant current, each additional transfer tank will reduce the current and voltage of each tank accordingly. If 2 or more transfer tanks need to be used at the same time, it is recommended to use a high power supply with high current, or set a lower current or voltage for a longer transfer time.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. Instructions for Use ： 1. Prepare the transfer buffer according to the table below.ReagentReduced SamplesNon-Reduced Samples Transfer Buffer (Powder, for Bis-Tris Gels)1 bottle1 bottle Antioxidant (400X) 1ml-Ethanol or Methanol100-200ml100-200mlH2OTo 1000mlTo 1000mlTotal Volume1000ml1000mlNote: Better transfer result can be obtained when using the ice-cold transfer buffer for the transfer. Unused transfer buffer can be stored at room temperature or 4℃ for up to two weeks, but should be discarded when its color turns light brown or yellowish brown.2. Transfer conditions: Protein transfer can be completed in one hour at 300mA constant current or 100V constant voltage (external ice bath is recommended). The transfer time can be appropriately reduced for smaller molecular weight proteins or increased for larger proteins. This transfer buffer is also suitable for rapid transfer, such as 500mA constant current for 15 minutes.3. For detailed Western procedures, please refer to http://www.aladdin-e.com/support/western.htm.