Antibody Sequencing Service

Antibody sequencing is a technology that precisely determines the full sequence of an antibody through high-resolution mass spectrometry and bioinformatics analysis. It covers the complete sequence from the N-terminus to the C-terminus, providing detailed information on both variable and constant regions. This technology identifies post-translational modifications (e.g., glycosylation) and disulfide bond linkages, offering a sequence foundation for antibody function studies and structural optimization.  

 

Antibody sequencing addresses key issues in antibody drug development, engineering modifications, and patent protection. The process typically involves sample preparation, protein digestion, mass spectrometry analysis, sequence data processing, and validation. Through antibody sequencing, researchers can define antibody sequences, ensuring stability and functional consistency in downstream applications.  

 



Figure 1. The Workflow of Antibody Sequencing

 

Services at MtoZ Biolabs

MtoZ Biolabs, an integrated chromatography and mass spectrometry (MS) services provider, provides advanced proteomics, metabolomics, and biopharmaceutical analysis services to researchers in biochemistry, biotechnology, and biopharmaceutical fields. MtoZ Biolabs offers antibody sequencing service leveraging advanced high-resolution mass spectrometry platforms and sophisticated bioinformatics technologies, handled entirely by an experienced research team. From sample preparation and protein digestion to mass spectrometry analysis and data interpretation, we provide a one-stop solution to ensure precision and efficiency in sequence determination. With extensive project experience and exceptional technical expertise, we are equipped to deliver complete sequence analysis for various antibody types, providing reliable support for your research needs. Our antibody sequencing service includes but is not limited to the following:

1. Antibody Sequencing Service

2. De Novo Antibody Sequencing Service

3. PCR Based Antibody Sequencing Service

4. Hybridoma Antibody Sequencing Service

 

If you are interested in our antibody sequencing service, please contact us freely.

 

Service Advantages

1. Comprehensive Antibody Sequence Analysis  

Optimized high-resolution mass spectrometry workflows and advanced de novo sequencing software ensure complete sequencing of the entire antibody sequence from the N-terminus to the C-terminus.  

 

2. Wide Applicability  

This technology is suitable for all antibody types (IgG, IgA, IgM, IgY, Fab, ScFv, etc.) and species (human, mouse, rat, rabbit, hamster, etc.). MtoZ Biolabs can also sequence conjugated or contaminated proteins, as well as antibodies with two different light chains. For murine and human antibodies, sequence analysis ensures the synthesized antibody retains its activity.  

 

3. High Throughput  

MtoZ Biolabs' antibody sequencing service leverages mass spectrometry-based sequence analysis with optimized workflows and bioinformatics processes. Whether sequencing a single antibody or hundreds, MtoZ Biolabs can efficiently meet your needs.

 

Case Study

1. Accurate Sequence Analysis of a Monoclonal Antibody by Top-Down and Middle-Down Orbitrap Mass Spectrometry Applying Multiple Ion Activation Techniques

This study highlights the use of top-down (TD) and middle-down (MD) mass spectrometry (MS) for accurate sequence analysis of monoclonal antibodies, minimizing sample manipulation to reduce artifactual modifications. Using an Orbitrap mass spectrometer, various ion activation methods, including ultraviolet photodissociation (UVPD) and high-capacity electron-transfer dissociation (ETD HD), were applied. The combination of 213 nm UVPD and ETD HD achieved a record sequence coverage of ~40% for intact antibodies, while MD strategies yielded up to 90% sequence information. The study also introduced the TDValidator software for systematic validation, optimizing data accuracy and enhancing confidence in MS2 spectra assignment. The antibody sequencing service applies advanced top-down and middle-down MS techniques to deliver detailed monoclonal antibody sequence analysis. Combining cutting-edge ion activation methods and rigorous data validation, this service ensures high sequence coverage and reliable characterization for complex antibody samples.

 



Fornelli, L. et al. Anal Chem. 2018.

Figure 2. Rituximab Sequence Analysis by Middle-Down Mass Spectrometry

 

2. Direct Determination of Antibody Chain Pairing by Top-down and Middle-down Mass Spectrometry Using Electron Capture Dissociation and Ultraviolet Photodissociation  

This study explores a method for directly analyzing antibody chain pairing using top-down and middle-down mass spectrometry combined with electron capture dissociation (ECD) and ultraviolet photodissociation (UVPD). The research demonstrates that this approach effectively deciphers the structural information of light and heavy chains while identifying disulfide bond linkages, offering a new tool for antibody structure research. The antibody sequencing service employs advanced mass spectrometry techniques to investigate antibody chains and structural characteristics, facilitating detailed chain pairing and structural analysis.

 



Shaw, JB. et al. Anal Chem. 2020. 

Figure 3.  The Combination of ECD and HCD Reveals Sequence Coverage Produced by the Light chain and Glycosylated Heavy Chain of Trastuzumab

 

Applications

1. New Drug Development: Used for developing antibody-based drugs by identifying sequence information to optimize drug design and functionality validation.  

 

2. Antibody Engineering: Providing precise sequence data for antibody modifications, affinity maturation, and humanization.  

 

3. Quality Control: Antibody sequencing service can assist in verifying antibody sequence consistency to ensure stability and reproducibility during production processes.  

 

FAQ

Q1: What are the key challenges in antibody sequencing using mass spectrometry?

Answer: Key challenges in antibody sequencing include the complexity of variable regions leading to incomplete sequence coverage, signal interference from multiple ion fragments after antibody digestion, and specificity limitations in distinguishing similar sequences. To overcome these issues, optimizing sample preparation to improve enzymatic digestion efficiency, using multiple ionization methods (such as ESI and MALDI) to enhance signal intensity, and employing high-resolution mass spectrometry combined with advanced data search algorithms can enable accurate antibody sequence determination with higher sensitivity. Our antibody sequencing service utilizes advanced mass spectrometry techniques and optimized workflows to tackle challenges in analyzing variable regions and resolving complex sequence ambiguities.

 

Deliverables

1. Comprehensive Experimental Details

2. Materials, Instruments, and Methods

3. Relevant Liquid Chromatography and Mass Spectrometry Parameters

4. The Detailed Information of Antibody Sequencing 

5. Mass Spectrometry Image

6. Raw Data