阿拉丁生产的UltraBio™ Western溶液套装（封闭液、洗涤液、一抗和二抗稀释液），是一种包含了Western Blot实验过程中常用的最新一代的快速高效溶液的套装。封闭液、一抗和二抗稀释液优于传统的同类产品，具有使用便捷、兼容性好、背景极低等优点。
本产品中的UltraBio™ WB高效一抗稀释液（无蛋白源）是最新一代的高效一抗稀释液，总体效果显著优于传统的基于BSA(牛血清蛋白)、脱脂奶粉等的一抗稀释液，可以用于Western Blot (WB)时一抗的稀释和配制。一抗稀释液具有以下优点：a.节约一抗，用本稀释液稀释和配制的一抗可以在4℃保存和使用不少于6个月，并且可以反复多次用于WB，利于节约一抗；b.背景极低，本稀释液不含血清和白蛋白，确保极高的信噪比；c.兼容性好，本稀释液后续兼容辣根过氧化物酶(Horseradish peroxidase, HRP)、碱性磷酸酶和生物素标记的二抗。本稀释液不含生物素，不会干扰基于生物素的检测；d.使用便捷，本稀释液无需添加任何额外的试剂，可以直接用于一抗的稀释。
本产品中的无蛋白快速封闭液是最新一代的快速高效Western封闭液，总体效果显著优于传统的基于BSA(牛血清蛋白)、脱脂奶粉、酪蛋白(Casein)等的封闭液及国外同类产品，主要用于Western blot (WB)实验中PVDF膜或硝酸纤维素膜(nitrocellulose membrane, NC膜)的封闭，也可以用于Western实验中一抗或二抗的稀释。无蛋白快速封闭液具有以下优点：a.快速高效，使用本封闭液时，封闭时间通常仅需5-15分钟，并且和BSA、脱脂奶粉、酪蛋白等传统的Western封闭液以及国外同类的快速封闭液相比，显示出更强的信噪比；b.背景极低，本封闭液不含血清和白蛋白，确保极高的信噪比；c.兼容性好，本封闭液兼容辣根过氧化物酶(Horseradish peroxidase, HRP)、碱性磷酸酶和生物素标记的二抗。本封闭液中添加了不影响HRP和AP活性的防腐剂，不会干扰HRP或AP标记二抗的检测。同时本封闭液不含生物素，不会干扰基于生物素的检测；d.使用便捷。本封闭液无需添加任何额外的试剂，可以直接用于印迹膜的封闭。关于不同封闭液的比较和选择，请参考阿拉丁的相关网页：http://www.aladdin-e.com/。
本产品中的Western洗涤液(10X)可以用于Western时一抗或二抗孵育后的洗涤。适当的洗涤可以降低背景，增强信噪比。
本产品中的UltraBio™ 无蛋白WB高效二抗稀释液是最新一代的高效二抗稀释液，总体效果显著优于传统的基于BSA(牛血清蛋白)、脱脂奶粉等的二抗稀释液，可以用于Western blot (WB)时二抗的稀释和配制。二抗稀释液具有以下优点：a.节约二抗。经本二抗稀释液稀释的二抗可以在2-3周内重复使用3-5次；b.背景极低。本稀释液不含血清和白蛋白，确保极高的信噪比；c.兼容性好。本稀释液兼容辣根过氧化物酶(Horseradish peroxidase, HRP) 、碱性磷酸酶和生物素标记的二抗。本稀释液中添加了不影响HRP和AP活性的防腐剂，不会干扰HRP或AP标记二抗的检测。同时本封闭液不含生物素，不会干扰基于生物素的检测；d.使用便捷。本稀释液无需添加任何额外的试剂，可以直接用于二抗的稀释。
按照每次Western blot需要封闭液10ml、一抗稀释液10ml、二抗稀释液10ml、洗涤液10ml+30ml+30ml+10ml=80ml，约100ml计算，一个包装的本试剂盒，大约可以进行10张膜的Western Blot检测。
组分和说明
使用说明
1. UltraBio™ WB高效一抗稀释液（无蛋白源）
a. 参考所用的一抗的说明，以及样品中目的蛋白的含量，按照适当比例例如1：1000、1：500等比例稀释一抗。一抗稀释后即可直接用于Western。一次Western结束后，可以回收稀释的一抗，4℃保存，以用于下次的Western。
2. 无蛋白快速封闭液
a. 完成转膜后，用Western洗涤液洗涤蛋白酶1-2分钟
b. 根据膜的大小，在平皿或者其它适当容器中倒入一定体积的封闭液，确保封闭液后续能充分覆盖膜即可。对于常规的western，一张约6.6×8.5cm的膜推荐使用约10ml左右的封闭液。
c. 用平头镊夹住膜的一角，将膜放置在 无蛋白快速封闭液中，使封闭液完全浸没膜，置于水平摇床上封闭约10分钟(通常5-15分钟均可；经多种抗体的测试封闭10分总的效果很多时候会显著优于常规的BSA封闭1小时的效果)。
3. ELISA&Western Blot 洗涤液
a. 取一瓶 ELISA&Western Blot 洗涤液，倒入到一洁净的量筒中，加入蒸馏水混匀并定容到1升后即为ELISA&Western Blot 洗涤液 (1×)，并可以用于Western的洗涤。没有用完的洗涤液可以室温保存，通常1-2周内是可以正常使用的。如果发现洗涤液出现浑浊、沉淀等异常现象，宜废弃。
b. 在一抗或二抗孵育结束后，加入少量 ELISA&Western Blot 洗涤液 (1×)，使之能盖住蛋白膜，在摇床上洗涤5分钟后，吸尽洗涤液，再加入新的洗涤液洗涤。共洗涤3次，每次5分钟。然后进行后续的操作。如果按照上述洗涤条件Western结果背景仍然较高，可以适当延长洗涤时间，并增加洗涤次数。通常，延长洗涤时间或增加洗涤次数不会对Western结果产生负面影响。
4. UltraBio™ Western二抗稀释液
a. 参考所用的二抗的使用说明，以及一抗的质量和样品中目的蛋白的含量，按照适当比例例如1：1000、1：500等比例稀释二抗。二抗稀释后即可直接用Western。一次Western结束后，可以回收稀释的二抗，4℃保存，以用于下次的Western。
b. 详细的Western操作可以参考我们相关网页：http://www.aladdin-e.com/support/western.htm
注意事项
1. 取放PVDF膜和NC膜应使用平头镊子，并仅轻轻夹取其边角，操作过程须避免膜表面产生划痕、折痕或压痕等痕迹。PVDF膜一经浸润和活化，需一直保持湿润，根据Western进行到的具体步骤可放置于western转模液或洗涤液等适当溶液中，否则可能会产生难以封闭的异常背景。
2. 为了能使稀释的一抗可以反复多次使用，一抗孵育结束后稀释的一抗应立即存放在4℃，以便于后续重复使用。
3. 通常无蛋白快速封闭液用于PVDF膜及NC膜时的封闭时间为5-15分钟。对于一些背景非常高的抗体，可以尝试将封闭时间延长为30-60分钟。此外，如有特殊需要，也完全可以4℃封闭过夜。由于没有任何一种封闭液是适用于所有实验体系的，因此对于一些特殊的实验，可能需要根据具体情况考虑使用其它更合适的封闭液。
4. 为了能使稀释的二抗可以反复多次使用，二抗孵育结束后稀释的二抗应立即存放在4℃，以便于后续重复使用。
5. 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。
6. 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™ Western Solution Kit (including Blocking Buffer, Washing Buffer, Primary Antibody Diluent, and Secondary Antibody Diluent) is a set containing the latest generation of fast and high-efficiency solutions commonly used in Western Blot experiments. The Blocking Buffer, Primary Antibody Diluent, and Secondary Antibody Diluent outperform traditional similar products, featuring advantages such as convenient use, good compatibility, and extremely low background.
UltraBio™ Primary Antibody Dilution Buffer
It is the latest generation of high-efficiency primary antibody diluents. Its overall performance is significantly superior to traditional primary antibody diluents based on BSA (Bovine Serum Albumin), non-fat milk powder, etc. It can be used for the dilution and preparation of primary antibodies in Western Blot (WB) experiments. It has the following advantages:
a. Primary antibody saving: Primary antibodies diluted and prepared with this diluent can be stored and used at 4°C for no less than 6 months, and can be reused multiple times in WB experiments, which is conducive to saving primary antibodies.
b. Extremely low background: This diluent does not contain serum or albumin, ensuring an extremely high signal-to-noise ratio.
c. Grate compatibility: This diluent is compatible with secondary antibodies labeled with Horseradish Peroxidase (HRP), Alkaline Phosphatase, and Biotin in subsequent steps. It does not contain biotin and will not interfere with biotin-based detection.
d. Convenient use: No additional reagents need to be added to this diluent, and it can be directly used for diluting primary antibodies.
Protein-Free Rapid Blocking Buffer
It is the latest generation of fast and high-efficiency Western blocking buffers. Its overall performance is significantly better than traditional blocking buffers based on BSA (Bovine Serum Albumin), non-fat milk powder, Casein, etc., as well as similar foreign products. It is mainly used for blocking PVDF membranes or Nitrocellulose Membranes (NC Membranes) in Western Blot (WB) experiments, and can also be used for diluting primary or secondary antibodies in Western experiments. It has the following advantages:
a. Fast and efficient: When using this blocking buffer, the blocking time usually only takes 5-15 minutes. Compared with traditional Western blocking buffers such as BSA, non-fat milk powder, and casein, as well as similar fast blocking buffers from foreign countries, it exhibits a stronger signal-to-noise ratio.
b. Extremely low background: This blocking buffer does not contain serum or albumin, ensuring an extremely high signal-to-noise ratio.
c. Grate compatibility: It is compatible with secondary antibodies labeled with Horseradish Peroxidase (HRP), Alkaline Phosphatase, and Biotin. A preservative that does not affect the activity of HRP and AP is added to this blocking buffer, which will not interfere with the detection of HRP or AP-labeled secondary antibodies. At the same time, it does not contain biotin and will not interfere with biotin-based detection.
d. Convenient use: No additional reagents need to be added to this blocking buffer, and it can be directly used for blocking blotting membranes. For the comparison and selection of different blocking buffers, please refer to Aladdin's relevant webpage: http://www.aladdin-e.com/.
ELISA&Western Blot Washing Buffer
It can be used for washing after the incubation of primary or secondary antibodies in Western experiments. Proper washing can reduce the background and enhance the signal-to-noise ratio.
UltraBio™ Protein-Free Secondary Antibody Dilution Buffer for Western Blot
It is the latest generation of high-efficiency secondary antibody diluents. Its overall performance is significantly superior to traditional secondary antibody diluents based on BSA (Bovine Serum Albumin), non-fat milk powder, etc. It can be used for the dilution and preparation of secondary antibodies in Western Blot (WB) experiments. It has the following advantages:
a. Secondary antibody saving: Secondary antibodies diluted with this secondary antibody diluent can be reused 3-5 times within 2-3 weeks.
b. Extremely low background: This diluent does not contain serum or albumin, ensuring an extremely high signal-to-noise ratio.
c. Grate compatibility: It is compatible with secondary antibodies labeled with Horseradish Peroxidase (HRP), Alkaline Phosphatase, and Biotin. A preservative that does not affect the activity of HRP and AP is added to this diluent, which will not interfere with the detection of HRP or AP-labeled secondary antibodies. At the same time, this blocking buffer does not contain biotin and will not interfere with biotin-based detection.
d. Convenient use: No additional reagents need to be added to this diluent, and it can be directly used for diluting secondary antibodies.
Recommended Usage Volume and Kit Capacity
Based on the calculation that each Western Blot experiment requires 10 ml of Blocking Buffer, 10 ml of Primary Antibody Diluent, 10 ml of Secondary Antibody Diluent, and 80 ml of Washing Buffer (10 ml + 30 ml + 30 ml + 10 ml), totaling approximately 100 ml of solutions, one package of this kit can be used for Western Blot detection of approximately 10 membranes.
Components
| W751600 | Component | 4×100ml | Storage | | W751600A | UltraBio™ Primary Antibody Dilution Buffer | 100ml | 2-8℃ | | W751600B | UltraBio™ Protein-Free Secondary Antibody Dilution Buffer for Western Blot | 100ml | 2-8℃ | | W751600C | Protein-Free Rapid Blocking Buffer | 100ml | 2-8℃ | | W751600D | ELISA&Western Blot Washing Buffer | 100ml | 2-8℃ |
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Instructions for Use
1. UltraBio™ Primary Antibody Dilution Buffer
a. Refer to the instructions of the primary antibody used and the content of the target protein in the sample, then dilute the primary antibody at an appropriate ratio (e.g., 1:1000, 1:500). The diluted primary antibody can be directly used for Western Blot (WB) experiments. After one WB experiment, the diluted primary antibody can be recovered and stored at 4°C for use in subsequent WB experiments.
2. Protein-Free Rapid Blocking Buffer
a. After completing membrane transfer, wash the protein membrane with Western Washing Buffer for 1–2 minutes.
b. Depending on the size of the membrane, pour a certain volume of Blocking Buffer into a petri dish or other suitable container, ensuring that the Blocking Buffer can fully cover the membrane afterward. For a regular Western Blot experiment, approximately 10 ml of Blocking Buffer is recommended for a membrane of about 6.6×8.5 cm.
c. Use flat-tipped forceps to hold one corner of the membrane, place the membrane in the Protein-Free Rapid Blocking Buffer (ensuring the membrane is completely immersed), and incubate on a horizontal shaker for about 10 minutes (usually 5–15 minutes is acceptable). Tests with multiple antibodies have shown that a 10-minute blocking time often yields significantly better results than the conventional 1-hour blocking with BSA.
3. ELISA & Western Blot Washing Buffer
a. Take one bottle of ELISA & Western Blot Washing Buffer (10×), pour it into a clean graduated cylinder, add distilled water, mix well, and dilute to a final volume of 1 liter to prepare ELISA & Western Blot Washing Buffer (1×), which can be used for washing in WB experiments. Unused Washing Buffer can be stored at room temperature and is generally usable within 1–2 weeks. If the Washing Buffer becomes turbid, develops precipitates, or shows other abnormalities, it should be discarded.
b. After the incubation of the primary or secondary antibody, add a small amount of ELISA & Western Blot Washing Buffer (1×) to cover the protein membrane. Wash on a shaker for 5 minutes, then aspirate all the Washing Buffer and add fresh Washing Buffer for another wash. Repeat the washing process 3 times, with each wash lasting 5 minutes, before proceeding to subsequent steps. If the background of the WB result is still high under the above washing conditions, the washing time can be appropriately extended and the number of washes increased. Generally, extending the washing time or increasing the number of washes will not have a negative impact on the WB results.
4. UltraBio™ Protein-Free Secondary Antibody Dilution Buffer for Western Blot
a. Refer to the instructions of the secondary antibody used, as well as the quality of the primary antibody and the content of the target protein in the sample, then dilute the secondary antibody at an appropriate ratio (e.g., 1:1000, 1:500). The diluted secondary antibody can be directly used for WB experiments. After one WB experiment, the diluted secondary antibody can be recovered and stored at 4°C for use in subsequent WB experiments.
b. For detailed Western Blot operation procedures, please refer to our relevant webpage: http://www.aladdin-e.com/support/western.htm
Precautions：
1. Use flat-tip tweezers to gently handle PVDF and NC membranes on the corners or edges to avoid producing any scrapes, creases or indentations on the surface of membrane. PVDF membrane, once infiltrated and activated, should be kept wet by placing the membrane in Western transfer buffer or washing buffer according to the Western Blot procedures. Otherwise, an abnormal background that is hard to be blocked may be produced.
2. To enable the diluted antibodies to be reused multiple times, the primary/secondary antibody dilutions should be stored at 4ºC immediately after each use.
3. Usually, it takes 5-15 minutes at room temperature to block PVDF or NC membranes. For some antibodies with very high background, prolong the blocking time to 30-60 minutes at room temperature, or overnight at 4ºC if necessary. Because no blocking reagent is optimal for all experimental systems, a proper blocking buffer should be selected based on the special requirement of your experiment. provides a variety of blocking solutions that you may compare and select.
4. To allow the diluted secondary antibody to be reused multiple times, the diluted secondary antibody should be immediately stored at 4°C after the secondary antibody incubation is completed, facilitating subsequent repeated use.
5. This product is for R&D only. Not for drug, household, or other uses.
6. For your safety and health, please wear a lab coat and disposable gloves during the operation.
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