Alt text – The best antibody sequencing tools for research

Antibody sequencing plays a crucial role in research to ensure the development of therapeutic antibodies, bispecific, CAR-T, and T-cell engagers. Determining the sequence of amino acids in an unidentified antibody aid in optimizing function, identifying new antibody clones, and banking databases.

Different approaches are available for sequencing, and researchers need to decide when in the discovery timeline to obtain sequences, considering both cost and bandwidth. This article provides an overview of the best antibody sequencing tools to help researchers make informed decisions.

Continue scrolling to find out the best antibody sequencing tools for your research.

The Best Antibody Sequencing Tools for Research

Antibody sequences are key for protein engineering and characterization in medicinal development. Users of antibody reagents can employ sequence analysis and alignment to determine binding and cross-reactivity, which is a precondition for rational experiment design.

Several online antibody databases store various data, such as suppliers, usage information, and publication citations. Other datasets, such as sequences from antibody genes or proteins, give significantly more information for antibody amino acid sequence and characterization.

Researchers need to use an antibody sequence alignment tool or de novo protein sequencing to decode a genome sequence that has yet to be sequenced. Researchers can utilize one of the many antibody sequence databases to discover the sequence when the sequence is known.

Several antibody sequence tools available include:

1. IMGT

The IMGT, also known as the ImMunoGeneTics information system, is a comprehensive collection of information that acts as a global reference. It specializes in human and other vertebrate immunoglobulins (Ig), Major Histocompatibility Complex (MHC), and T cell receptors (TcR) molecules.

The IMGT database contains archives on antibody sequencing, genomic structure, and structure. For example, IMGT's monoclonal antibodies database (IMGT/mAb-DB) is a unique, peer-reviewed data source on mAbs (monoclonal antibodies) for clinical contexts and fusion proteins in immune-related research.

2. AbYsis

abYsis is an interactive antibody research platform that houses consolidated antibody sequencing and structural data collection.

AbYss has three primary uses. It may be used to do database searches to learn more about sequences (and structures), such as detailed annotations, the position of unusual residues, and post-translational modification sites.

It will also assist you in examining regular trends throughout the sequencing in the database and entering your sequences for additional study.

3. UniProt

UniProt is one of the most comprehensive, high-quality, and easily accessible resources for protein production, sequence, and functional information. Because antibodies are not UniProt's primary focus, the antibody sequence database only contains a representative group of germline immune response sequences and a small assortment of non-germline antibody sequences.

4. cAb-Rep

cAb-Rep is a database of how to find antibody sequence from well-chosen antibody repertoires. It comprises 306 B cell repertoires collected from 121 human donors, including individuals with healthy, vaccinated, and autoimmune illnesses.

The cAb-Rep database presently contains 267.9 million curated full-length V(D)J light chain transcripts and 72.9 million full-length V(D)J light chain transcripts produced by Illumina sequencing technology.

5. SabDab

The SabDab Structural Antibody Database (SAbDab) is a database that comprises all of the antibody structures presently accessible in the PDB. Furthermore, all of these structures are annotated and presented consistently.

The user can utilize these qualities, in addition to structural criteria such as complementarity defining area loop conformation and changing domain orientation, to focus the search for a suitable structure.

6. IgBLAST

IgBLAST matches immunoglobulin and T cell receptor sequences against a germline database to identify V(D)J recombination and somatic mutation status. It’s extensively utilized in academic and clinical settings to find novel genes and alleles.

Comparison of the Leading Antibody Sequencing Tools

Abysis is a web-based tool that allows users to analyze and compare antibody sequences from various sources, including IMGT and other databases. It offers a range of tools for sequence alignment, phylogenetic analysis, structure prediction, and visualization tools for exploring the properties of different antibodies.

Each of these tools offers unique features and benefits. IMGT is known for its comprehensive database and advanced analysis tools, while IgBLAST is highly accurate and reliable for identifying novel genes and alleles. Abysis is a versatile tool with various analysis and visualization options.

Factors to Consider When Choosing an Antibody Sequencing Tool

Antibodies have become indispensable tools in life sciences, diagnostic applications, and therapeutic drug delivery. However, selecting the right antibody software is crucial, as it can impact the experiment's accuracy, sensitivity, specificity, cost, and ease of use.

With over 4.5 million commercial antibodies available from 350+ suppliers, choosing the right one can be challenging. Fortunately, here are some factors to consider when selecting an antibody sequencing tool.

Accuracy

When choosing an antibody sequencing technology, accuracy is a key consideration. The capability to get accurate and trustworthy findings depends on the instrument's accuracy. The antibody must be very accurate in detecting the target protein and verified for the intended use.

Sensitivity

The instrument's sensitivity governs its capacity to identify minute quantities of the target protein. The custom antibody must be extremely sensitive to guarantee that even the tiniest quantity of the target protein can be identified.

Specificity

The instrument's capacity to discriminate between the target and other proteins is known as specificity. It should be peculiar to ensure the antibody does not cross-react with other proteins and produce unreliable findings.

Cost

You can weigh the price of the antibody sequencing technique against its accuracy, sensitivity, and specificity. The tool's performance must justify its expense.

What are Reliable Antibody Sequencing Services?

Acquiring intellectual property rights in medicinal research and development depends on antibody sequencing. To prevent contamination, genetic drift, gene loss or mutations, and hybridoma storage mishaps, an antibody must be sequenced with a 100% correct sequence.

It also offers a model for several downstream application platforms, including isotype/format switching, humanization, affinity maturation, and other possibilities for antibody creation.

Working with a professional and experienced antibody sequencing serviceprovider is essential to obtain reliable and accurate antibody sequencing analysis results. Several service providers offer comprehensive antibody sequencing services, including hybridoma and de novo antibody sequencing services.

De Novo Antibody Sequencing

De Novo Antibody Sequencing is a unique service that can sequence monoclonal antibodies (mAbs) even without hybridoma cell lines with 100% coverage of the targeted areas and exceptional accuracy.

In a 3-week timeframe, the amino acids in CDRs may be firmly mapped to over 20 distinct peptides and 100% of the sequence, and pairs of intense fragment ions securely support them in at least ten scans.

The database checks the mass spectrometry data obtained after conducting sequence prediction based on the mass spectrum (label CDR region). Only the precise match result is sent to the customer.

Hybridoma Sequencing

Hybridoma Sequencing necessitates sequencing more than five antibody-independent clones following RT-PCR and rapid amplification of cDNA ends (RACE) to avoid interference from pseudogenes. This process produces 100% accurate hybridoma sequencing analysis.

The Hybridoma Sequencing service covers a variety of animals, including mice, rats, rabbits, goats, and chickens. If humanization or other antibody engineering techniques are included in therapeutic antibody studies, they can start after the VL and VH sequences are supplied.

Protein and Peptide Synthesis Services vs Antibody Sequencing Service

Protein and peptide synthesis services produce specific polypeptide sequences essential for applications such as antibody sequencing, designing enzymes, testing drugs, gene synthesis, and creating antibodies. This section discusses the process of obtaining protein andpeptide synthesis services, examples of service providers, and the benefits and drawbacks of using these services.

Obtaining Protein and Peptide Synthesis Services

Rather than engaging in custom peptide synthesis, many scientists choose to outsource the job to specialized service providers. There are two main avenues of peptide synthesis: solid-phase and liquid-phase synthesis.

Attaching the C-terminus to a solid resin, which makes it easier to separate the peptide from the reaction mixture, is known as solid-phase synthesis and is increasingly often used.

In contrast, liquid-phase synthesis takes longer and requires more work. Still, it offers the benefits of several purification steps and the potential for convergent synthesis, in which smaller produced peptides combine to generate bigger ones.

Protein and Peptide Synthesis Service Providers

Several protein and peptide synthesis providers offer customizable options that include the addition of modifications such as phosphorylation, methylation, biotinylation, glycosylation, cyclization, or attachment to carrier proteins or dye labels, quantity, purification, and additional testing.

Benefits and Drawbacks of Using Protein and Peptide Synthesis Services

One of the main benefits of using protein and peptide synthesis services is the convenience and time-saving factor of outsourcing the production of custom peptides. Additionally, service providers offer a range of customizable options to tailor peptides to specific applications like the production of Elisa Kits.

However, a drawback of using these peptide synthesis services is the potential for higher costs than in-house production, especially for large-scale projects. It's also important to note that the quality of the recombinant protein productionmay vary depending on the recombinant antibody service provider, so choosing a reputable and reliable provider is essential.

Antibody Sequencing: Understanding the Facts and Benefits

Antibodies are proteins B-lymphocytes produce in the immune system in response to a specific antigen. These proteins attach to the antigen's binding site with excellent specificity and remove it from the bloodstream.

Generally, antibodies are essential tools in biotech and biosensor development. Their specificity has several applications, such as attaching them to sensor surfaces to detect antigens or using tagged detection platforms.

Antibodies can be produced directly by the body, but custom antibody synthesis offers a high purity level achieved using a 4mg immunogen. The custom antibodies are polyclonal or monoclonal, depending on how they are produced and extracted. Polyclonal antibodies recognize multiple epitopes on an antigen, while monoclonal antibodies remember only one antigen epitope.

In custom polyclonal production, animals like goats, rats, mice, and chickens need immunization before being examined to produce antigens. Rabbits are preferred in this process because they produce more antibodies than other animals due to their high responses to antigens, making it cost-effective.

Moreover, rats and mice are ideal when processing a small amount of antiserum. On the other hand, goats produce many antibodies, but they have a lower response rate to antigens.

How is Antibody Production Done?

The five-step technique of producing antibodies consists of prepping the antigen, immunizing hybridoma development, fusing hybridoma development, subcloning, and producing and purifying monoclonal antibodies.

Successful custom antibody production service necessitates meticulous preparation and execution of numerous critical procedures and considerations. There is also a need to purify or synthesize the objective protein, select an immunogenic carrier protein, and combine the antigen with the carrier protein to make an immunogen.

Antibody characterization is crucial after protein expression, especially in biopharmaceutical development and research, where diverse alterations occur throughout different synthesis phases.

The relative specificity of the antibody to the protein of interest needs simple Western analysis tracking for monoclonal antibody database and polyclonal antibody sequencing. Proteolytic enzymes can produce antibody fragments, resulting in smaller antigen-binding fragments.

Antibody Sequencing Tools for Research FAQs

What is antibody sequencing?

Antibody sequencing is the process of determining the nucleotide sequence of the variable regions of antibodies. It provides a comprehensive understanding of the antibody repertoire of an organism, including the diversity and distribution of antibody clones.

What are the benefits of antibody sequencing?

Antibody sequencing enables researchers to understand better antibody binding specificity and variety, which can assist in the development of more effective medicines and diagnostics. It also allows for identifying distinct antibody clones to generate monoclonal antibodies.

What are the challenges associated with antibody sequencing?

The complexity of the antibody repertoire is one of the key problems of antibody sequencing because it comprises millions of distinct antibody clones. Moreover, the significant variety of antibody sequences and the requirement for precise sequencing might be a technological hurdle.

Conclusion

Antibody sequencing is essential in various research areas, enabling scientists to develop molecular recognition tools with excellent specificity for detecting substances of interest. Custom antibody production requires high purity and meticulous preparation to obtain high-quality antibodies for various research activities. Antibody characterization is also essential in biopharmaceutical development and research to determine the specificity of the antibody to the protein of interest.