Creative Biolabs offers academic and industrial researches on the high resolution paratope mapping of antibody all over the world, depending on novel research methods and tools. We are capable of meeting the specific need of each customer with the best project design.

Background

Paratope mapping is characterization of the binding sites, or paratopes of antibodies which recognized by antigens. However, to develop vaccines and bio-therapeutics, one of the most important processes is identification of the antibody paratopes. There are a number of crucial uses to understand the paratope in the antibody, for example, predicting suitable antigens as vaccine components, improving our knowledge of the immune response and autoimmunity, gaining an supernormal understanding of a therapeutic antibody's mechanism of action.

Fig.1 Paratope Mapping using computational procedure (Xiang Fang et al.,2012).

Approaches of Paratope Mapping

Compared with conventional approaches, Creative Biolabs has developed a number of novel research methods to perform paratope mapping, which is lower cost and faster, including:

• Paratope mapping using computational procedure

Rigid body docking, free and steered molecular dynamics (MD) simulations and homology modeling constitute the computational procedure. With the computational procedure, it is reachable to identify key paratope residues on the target antibody and their partners on antigen. The computational model of quaternary structure of complexes consisted of more than two biological macromolecules which interact with each other is macromolecular docking. Molecular dynamics (MD) is an analysis to research the motion of molecules and atoms. To overview the dynamical evolution of this system, molecules and atoms are performed to interact for a certain time. Comparative modeling of protein, termed homology modeling as well, builds an atomic-resolution models of the protein from its amino acid sequence, and furthermore builds an experimental three-dimensional structure of a correlative isogenous protein.

• Paratope mapping using Hydrogen–deuterium exchange and mass spectrometry

By using Hydrogen-deuterium exchange and mass spectrometry, in the solvent hydrogen atoms will exchanged by backbone amide hydrogen atoms and if there is deuterium in the solvent deuterium but don't contains hydrogen then the same exchange process occurs so that the protein can be deuterated. The flexibility and the availability of the target part of the protein in the solvent decide the rate of any special hydrogen atom happens. The hydrophobic core of a rigid protein where hid hydrogens will exchange at all, while those on a flexible loop will exchange nearly instantaneously.

• Paratope mapping using Peptide-based approaches

Peptide-based approaches are based on the composition of overlap peptides, however the peptides must contain the entire sequence of the antigen. Following the peptides fixated onto a solid surface as an array and the binding to the antibody of curious is determined in an ELISA format. This approach can make antibodies 'panned' with very huge phage libraries available. Furthermore, the recombinant peptide genes from binding phage particles can be sequenced to identify the sequences and putative epitopes.

• Paratope mapping using Hopfield network

Using a Hopfield Network while representing the epitope and paratope in a complex as 3-dimensional attributed graphs, a mapping is estimated from a random training sample while representing. With the help of KBCM when using Hopfield Network, mapping paratopes on immunoglobulins to the epitopes can be available.

Creative Biolabs provides other various Epitope mapping services. Please feel free to contact usfor a detailed quote.

Other optional antibody analysis services:

• Antibody Characterization
• Antibody Affinity Measurement
• Hybridoma Sequencing Service
• De Novo Antibody Sequencing Services
• Monoclonal Antibody Epitope Binning
• SPR Services
• Isothermal Titration Calorimetry (ITC) Services
• Differential Scanning Calorimetry (DSC) Services
• Dual-Polarization Interferometry (DPI) Services

Published Data

• Epitope and Paratope Mapping by HDX-MS to Elucidate the Difference of Two Anti-NadA MAbs

Fig. 2 Paratope mapping of NadA on 1C6 and 7F11. (Laura R Grauslund, 2021)

The characterization of the antigen-antibody interaction is very important to understand the function of antibodies. Bexsero is a multicomponent vaccine against Neisseria meningitidis. One of the key vaccine antigens is a trimeric coiled-coil protein, Neisseria adhesin A (NadA). Two NadA specific monoclonal antibodies (mAbs) were isolated from individuals vaccinated with Bexsero. Both of them had similar binding affinity, but only one kind of mAb had bactericidal activity. Here, the researchers studied the interaction between two mAbs and NadA antigens by using hydrogen/deuterium exchange mass spectrometry (HDX-MS) combined epitope and paratope mapping strategies. The epitope mapping confirmed that one of the antibodies binds to NadA, but the paratope mapping showed that both mAbs bind to NadA through several complementary determining regions across the heavy chain and light chain. This result highlights the advantage of HDX-MS assay combined epitope and paratope mapping strategy to characterize antigen-antibody interactions.

FAQ

1. What is paratope mapping of antibodies?

   Paratope mapping of antibodies refers to the process used to identify the specific region on an antibody that interacts with the antigen. This region, known as the paratope, is crucial for the antibody's ability to recognize and bind to a specific epitope on the antigen. Mapping the paratope helps in understanding the molecular basis of antibody-antigen interactions and is essential for antibody engineering and therapeutic development.

2. Why is paratope mapping important in antibody research?

   Because it provides detailed insights into the antibody's mechanism of action. By knowing exactly where and how an antibody binds to an antigen, researchers can design more effective antibody-based therapies, improve antibody specificity and affinity, and minimize off-target effects. This information is also critical for the development of vaccines and diagnostic tools.

3. What techniques can be used in paratope mapping?
   • X-ray Crystallography: Provides high-resolution details of the antibody-antigen complex.
   • Nuclear Magnetic Resonance (NMR) Spectroscopy: Useful for studying antibodies in solution.
   • Mutagenesis: Involves mutating specific amino acids in the antibody's variable region to assess changes in binding affinity.
   • Hydrogen/Deuterium Exchange Mass Spectrometry (HDX-MS): Measures the protection of peptide bonds from hydrogen to deuterium exchange upon antigen binding.
   • Epitope Mapping Technologies: Such as peptide scanning, phage display, or array-based techniques to determine interacting residues.
4. How does paratope mapping contribute to therapeutic antibody development?

   Paratope mapping allows researchers to identify and engineer antibodies with enhanced affinity and specificity for their target antigens. By understanding the binding interface, developers can modify antibodies to increase their effectiveness, reduce immunogenicity, and improve stability. This detailed mapping also facilitates the design of antibody-drug conjugates and bispecific antibodies where precise interaction with multiple targets is required.

5. Can paratope mapping be used to predict cross-reactivity of antibodies?

   Paratope mapping can help predict cross-reactivity by identifying the key residues in antibodies that interact with the epitope. By analyzing these interactions, researchers can determine whether an antibody might bind to similar epitopes on different antigens, which is particularly important in avoiding unintended interactions in therapeutic applications. This is critical for ensuring the safety and efficacy of antibodies used in clinical settings.

6. What role does computational biology play in paratope mapping?

   Computational biology plays a significant role in paratope mapping by using algorithms and modeling techniques to predict antibody-antigen interactions. These methods can include molecular docking, which predicts how antibodies and antigens fit together, and molecular dynamics simulations, which explore how these complexes behave over time. Computational approaches can significantly reduce the need for extensive experimental procedures by narrowing down the potential binding sites and guiding targeted mutagenesis experiments.

7. Are there any recent advancements in paratope mapping technologies?

   Recent advancements in paratope mapping technologies include the use of next-generation sequencing and deep learning to better predict and analyze antibody-antigen interactions. Techniques such as cryo-electron microscopy (cryo-EM) have also become more prevalent, allowing for the visualization of antibodies bound to antigens at near-atomic resolution without the need for crystallization. These technologies enhance the speed and accuracy of mapping and are opening new avenues for antibody research and development.

Resources

Use the resources in our library to help you understand your options and make critical decisions for your study.

All listed services and products are For Research Use Only. Do Not use in any diagnostic or therapeutic applications.