- It is necessary to develop a new research project to find out the basic characteristics of the protein of interest, especially if the structure of the protein is known, it will be helpful to select the recognition region which is easy to contact and recognize.
- However, the protein conformation will affect the interaction between the antibody and its recognition region.
- Continuous and discontinuous recognition region continuous region refers to the recognition region composed of continuous amino acid sequence (residue).
- In order to avoid the risk that the recognition region is hidden in the protein, we usually recommend selecting the N and C ends of the protein to produce the corresponding antibody.
It is necessary to develop a new research project to find out the basic characteristics of the protein of interest, especially if the structure of the protein is known, it will be helpful to select the recognition region which is easy to contact and recognize.
However, in the absence of such precise structural information (most of which is the case), understanding the purpose (application) of the research will affect the strategy of peptide design.
For example, if the research focuses on different regions of the protein, such as the C-terminal or N-terminal, or the protein in a specific state, such as phosphorylation, then the peptide designed according to the required sequence and the corresponding antibody produced should not be too difficult in application.
However, the protein conformation will affect the interaction between the antibody and its recognition region.
The possible problem in this case is that if the recognition region is hidden inside the protein in the folded protein, the antibody will not be able to reach the region.Generally speaking, the most ideal antigenicity recognition region should be hydrophilic, located on the surface of protein and easy to deform.
In most natural (NATURAL) environments, hydrophilic regions tend to concentrate on the protein surface, while hydrophobic regions are often wrapped in the protein interior.
Similarly, antibodies can only interact with the recognition regions found on the egg surface, and when these recognition regions have enough structural deformability and are transferred to the position where the antibodies can be contacted, there will be a high level of interaction between antibodies and these recognition regions Affinity.
Continuous and discontinuous recognition region continuous region refers to the recognition region composed of continuous amino acid sequence (residue).
Most of the antibodies are targeted at the continuous recognition region, and the antibody can bind to such regions with high affinity, indicating that the sequence is not in the protein.
The discontinuous recognition region represents a polypeptide sequence with a certain folding, or the recognition region of an antibody connecting two separated polypeptides.
In some cases, antibodies against such discontinuous recognition region can also be produced, but the antigenic peptide used for immunization must have a secondary structure similar to the discontinuous recognition region, and the length of the sequence needs to meet the relevant requirements.
In order to avoid the risk that the recognition region is hidden in the protein, we usually recommend selecting the N and C ends of the protein to produce the corresponding antibody.
Because in intact proteins, the ends of N and C are usually exposed to the protein surface. However, it must be noted that the C-terminal hydrophobicity of membrane protein is too strong to be used as antigen.
In general, we recommend that the sequence length of antigen polypeptide should be 8-20 If the amino acid residues are too short, there is a risk that the peptide is too special and the affinity (binding capacity) between the antibody and the natural protein is not strong enough.
Similarly, if the sequence length is more than 20, it may be introduced into the secondary structure, resulting in the possibility of losing specificity of the antibody. Moreover, the longer the peptide chain is, the more difficult the synthesis is, and it is not easy to obtain high-purity production Products.
The basic selection principle of carrier protein cross-linking: the carrier protein is added at the end far away from the antibody recognition region, and adding Cys at n or C terminal is the preferred method for cross-linking when there is no Cys in the sequence.
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