Essential guidance 2 | Fishing with Purekine “fishing gear”, Protein Association “wishers hooked”

 

 

In the last issue, the editor brought you “Dry Goods | Using IP “Hook” Fishing, Protein Club “Wishers Take the Bite””, and the response was very enthusiastic. In the voice of everyone, the editor is rushing through the previous experimental notes, bringing you the second bullet of protein interaction knowledge-Pull-Down. If the protein interaction is compared to an interesting “fishing” activity, then the Purekine filler we use is a tough and slender “fishing rod” and “fishing line”. The known GST-labeled recombinant protein is sharp The strong “fish hook” and “fish bait” are the perfect combination, and our target protein is the “fish” that moves freely in the water.

The concept of Pull-Down is similar to Co-IP, and its purpose is to study proteins or ligands that bind to known bait proteins. Pull-Down aims to prove the interaction between two proteins or explore unknown proteins or molecules that can bind to the target protein. Pull-Down is different from IP or Co-IP in that it is not based on antibody-antigen interaction, not an immune response. The bait protein (or ligand) is immobilized on a solid support through a non-antibody affinity system. This immobilization can be covalently coupled to the activated microbeads, or through the receptor molecule on the support The bound affinity tag binds and thus immobilizes. For example, immobilized metal chelating affinity chromatography (IMAC) resin can be used to pull-down histidine tag bait protein. The optimization of Pull-Down needs to consider the special nature of the affinity system used. Principle of Pull-Down According to the following diagram, you can understand the principle of Pull-Down:

Pull-Down

Coomassie brilliant blue

Comparison of Pull-Down and protein purification experiments:

  1. Use recombinant gene expression protein, and then perform downstream protein extraction and purification experiments;
  2. Analysis of the similarities and differences between the two experimental procedures;
Experiment type Pull-Down protein purification
Solid carrier Affinity chromatography resin Affinity chromatography resin filler
Carrier form In bulk Column
Incubation method Shaker incubation Incubate in the purification column
Type of buffer Binding Buffer

Wash Buffer

Elution Buffer

Binding Buffer

Wash Buffer

Elution Buffer

Target protein form Prey protein Recombinant protein
  1. Types of products to be tested:
Product Category Item No.
His tag protein purification tool BMR20000
GST tag protein purification tool BMR20100
MBP tag protein purification tool BMR20206
Flag tag protein purification tool BMR21504
Biotin tag protein purification tool BMR20306
Protein A Antibody purification tools BMR21604
Protein GAntibody purification tools BMR20600
Protein A/G Antibody purification tools BMR20704
Protein L Antibody purification tools BMR20800

Conclusion

The Purkine series of protein purification tools provided by Abbkine are multifunctional and suitable for a variety of protein experiments. When your target protein is a recombinant protein, the purification medium can be used for the extraction of recombinant protein, which is a protein purification experiment; when your target protein is a prey protein that interacts with the recombinant protein, this series of products can be used for prey Extraction and purification.

This issue mainly talks about the knowledge of Pull-Down, time is limited, I have to say goodbye to everyone. At the same time in the next issue, we will have exciting content to meet you all. Stay tuned~~~~~

You are welcome to pay more attention to Abbkine’s official account, and there will be more scientific research surprises waiting for you. At the same time, in order to provide readers with a platform for learning biological knowledge, Abbkine’s technical experts have worked tirelessly to sort out the dry goods of a variety of high-end biological experiments, which will be published in the WeChat public account in the later period. Please continue to pay attention to Abbkine for readers who love science. No public.

Abbkine focuses on the fields of proteinology and cytology, and is committed to the innovation and research and development of various antibodies, proteins, analytical reagents and kits, in order to become a key promoter in the development of life science research, drug development and other fields. We provide you with the favorite products of protein and immune research users, from basic immunological products, such as protein extraction and quantification, to internal reference label antibodies, primary and secondary antibodies for immunological experiments; the favorite products of cell research users, from Dyes and kits for detecting cell status, organelle extraction kits, cell substructure staining and tracking and cell metabolism detection products, to cytokine and protein detection kits for cell culture, just to help your research career !

发表评论

电子邮件地址不会被公开。 必填项已用*标注