Monoclonal Antibody Development

BioGenes has a proven track record with more than 1,000 successfully completed monoclonal antibody (mAb) projects that resulted in the generation of highly specific and sensitive diagnostic reagents and their application in multiple R&D ELISA kits. With over 20 years of experience BioGenes possesses profound technical and analytical know-how and capabilities in the generation of highly pure and consistent antibodies at scalable amounts. Customized screening systems are implemented for the selection of the right clones against every antigen.

The hybridoma technology allows for the generation of genetically highly diverse populations of monoclonal antibody secreting cell lines. Our immunization strategies are designed to fully exploit the positive effects of in vivo affinity maturation of the antibodies to be generated and, thus, facilitate the development of highly specific and sensitive immunoassay reagents. Furthermore, our workflows ensure the provision of highly reliable and reproducible reagents for a broad spectrum of customer applications. 

New in 2024

Our full range of services has been expanded to include antibody sequencing.

In addition, our in-house animal facilities and laboratories are now equipped to safety levels 1 and 2 (S1, S2) in accordance with the German Genetic Engineering Act (GenTG/GenTSV) and to risk levels 1-3** in accordance with the German Biological Substances Ordinance (BioStoffV). We now also accept corresponding antigens.

Development Strategy

Each project is conducted in close cooperation with the customer and consists of different work phases, each of which can be adapted according to the project’s specific aims. 

Phase 1 - Expert consultation

  • Definition of the project’s aims and challenges
  • Thorough discussion of the antigen characteristics 
  • Support in antigen design 
  • Definition of screening set-up (inhibition tests and/or screenings of cross-reactants)

Phase 2 - Immunization

  • Antigen generation (peptide synthesis or antigen conjugation)
  • Antigen characterization
  • Immunization of mice according to BioGenes’ enhanced immunization protocol or customized protocols (upon request)
  • Titre determination and selection of mice for hybridoma generation

  • Optional: Prolongation of immunization schedule to achieve appropriate titres


Phase 3 - Cell fusion & hybridoma screening

  • Generation and selection of hybridoma cell lines
  • Screening for IgG producing hybridomas by ELISA


  • Testing of multiple cross-reactants for antibody characterization
  • Provision of spleen cells for customer processing 
  • Delivery of hybridoma cell suspensions for testing by the customer

Phase 4 - Cloning

  • At least two cloning cycles and screening for relevant hybridomas
  • Delivery of cell culture supernatants for each clone for in-house testing by the customer
  • Characterization of selected clones (isotype determination and mycoplasma testing)


  • Adaption to serum-free medium
  • Pair Search – antibodies will be biotinylated and tested in an ELISA to identify suitable capture and detector antibodies

Phase 5 - Delivery & preservation

  • Cryo preservation of the relevant hybridomas
  • Delivery of three cryo vials per clone

Approximate project duration: 5-8 months

Additional Services


Antibody sequencing/hybridoma sequencing allows to determine the variable antibody gene sequences quickly and efficiently conferring the specificity of the antibody. This opens new possibilities in the field of recombinant antibody technologies and permanently establishes antibody identity. Due to our 30 years of experience in antibody services and our commitment to provide our customers with a complete service package we have now added antibody sequencing to our portfolio. Ask us for a quote at any time.


Test Production

We recommend a test production if there is insufficient information on the production rate of a clone of interest or if you need antibody amounts of 50 mg or more. The test production is based on the affinity purification of the antibody from 350 ml cell culture supernatant. 


Large-Scale Antibody Production 

BioGenes offers the bulk production of monoclonal antibodies at various scales and quantities ranging from research (mg) to commercial (g) scale amounts per batch. The production is carried out with hybridoma cells specifically developed by BioGenes or with cells provided by the customer. 


Longterm Storage and Maintenance

Cell viability and the rate of antibody production from established hybridoma cell lines can diminish over time. In many cases, the antibody production can be restored by re-cloning the hybridoma cell line. The hybridoma re-cloning and development can be ordered separately, if necessary.

  • Routine maintenance once every two years (or other time intervals as requested by the customer)
  • Includes thawing, re-cultivation and additional cloning
  • Screening for high productivity and preparation of new vials for cryo conservation


If you need more information, a specific offer 
or want to talk to an expert


What is the definition of hybridomas in antibody discovery?

Hybridomas are immortal antibody-secreting cell lines developed by Georges Kohler and Cesar Milstein in 1975. Created by fusing short-lived plasma cells from pre-immunized hosts with myeloma cells, hybridomas offer long-lasting production of high-quality monoclonal antibodies.

What are the advantages of using hybridomas in antibody discovery?

Hybridomas offer natural advantages, such as preserving VH/VL pairing, efficient in vivo affinity maturation, and the ability to produce full-length IgG antibodies without additional sequencing or cloning. These features make hybridomas invaluable for antibody discovery in therapy, research, and diagnostics.

Which animals are utilized for the development process?

BioGenes employs mice of the strains BALB/cJRj, RjOrl:SWISS, and C57BL/6NRj, as well as rats of the strain RjHan:SD - Sprague Dawley for immunization. These strains possess a high repertoire of antibody genes. The genetic foundations of antibody production in humans and mice are biologically very similar compared to other animal species. Furthermore, the required fusion partners and techniques, which are patent-free, are exclusively based on the mouse system.



What are the major uses of hybridoma cell lines?

After proper screening and monoclonal selection, mouse hybridomas secrete large quantities of IgG monoclonal antibodies. These antibodies can be conjugated to enzymes or fluorescent labels for specific antigen detection or capture. The versatility of hybridomas is particularly beneficial for diagnostics and research, aiding in disease detection and unraveling underlying causes.

Why does BioGenes use hybridoma technology instead of phage display for antibody development?

  1. Proven Track Record: Hybridoma technology has a well-established track record and remains a proven method for generating monoclonal antibodies (mAbs). With over 20 years of experience, BioGenes has successfully completed over 1,000 mAb projects, demonstrating the effectiveness of this technology.
  2. Specificity and Sensitivity: Hybridoma technology allows for the generation of highly specific and sensitive antibodies. This is crucial for applications in diagnostics and research where precise results are required.
  3. In Vivo Affinity Maturation: BioGenes' immunization strategies are designed to harness the positive effects of in vivo affinity maturation of antibodies. This leads to the development of highly specific and sensitive immunoassay reagents.

What challenges do hybridomas face in the long term?

Hybridomas may suffer from instability due to mutations or chromosome loss over time. To address this, hybridoma cell lines can be sequenced, enabling adaptation of antibody-encoding genes to recombinant expression systems for improved stability.

How does the hybridoma selection process work?

The hybridoma selection process involves separating a mixture of unfused and fused cells generated during the fusion of plasma cells with myeloma partners. Plasma cells, with short lifespans, are easily eliminated, while unfused myelomas are selectively removed by utilizing myeloma partners lacking HGPRT. The subsequent culture in HAT medium allows only cells with the HGPRT enzyme (hybridomas) to survive, ensuring the isolation of successfully fused cells for sustained antibody production.

How are hybridomas screened?

Hybridomas are commonly screened using the Enzyme-Linked Immunosorbent Assay (ELISA). During ELISA screening, the supernatant from each hybridoma is tested for its ability to bind to a specific antigen. Positive hybridomas, which successfully produce antibodies targeting the antigen, are then selected for further characterization and development.

Why are positive hybridoma cells subcloned?

After identifying a positive well through initial ELISA screening, hybridomas are transferred to larger volumes (e.g., 24-well plates) in preparation for subcloning. This process, often conducted via the limiting dilution method, ensures the isolation of stable monoclones. By diluting hybridoma cultures and dispersing them into 96-well plates, monoclonality (one cell per well) is achieved. Performing limiting dilution at least two times helps reduce the risks of producing mixed hybridoma populations, ensuring the purity and clonality of selected hybridoma cell lines for consistent antibody production.

What is the ideal antigen for monoclonal antibody production?

The key to successful hybridoma generation lies in the host's immune response, necessitating immunogenic antigens for effective immunization. The choice of antigens, such as proteins, peptides, or haptens, plays a critical role. Proteins and peptides, preferred for precise targeting, enable antibodies from hybridomas to focus on specific regions of pathogens or protein complexes. While peptides may lack sufficient immunogenicity, their conjugation with carriers (adjuvants) enhances recognition by the host organism. To obtain more information about our antigen requirements please click here.

What are the primary applications of hybridoma-generated antibodies?

Murine antibodies are well-suited for diagnostic applications. The cost efficiency of mouse hybridoma technology and the ability to produce small quantities of monoclonal antibodies through hybridomas make them suitable for diagnostic purposes. To make them suitable for therapeutic applications, murine antibodies often undergo humanization, a process that extends the turnaround time but is essential for minimizing adverse reactions and enhancing therapeutic efficiency.

Is it feasible to use a single monoclonal antibody across various applications?

Developing an antibody that performs effectively across diverse applications like flow cytometry, ELISA, Western Blot, Immunohistochemistry, etc., is highly challenging. The variation in assay conditions and sample preparation methods among these platforms poses a significant hurdle. For example, Western Blot applications require antibodies targeting linear epitopes (peptides) due to protein denaturation, whereas flow cytometry relies on maintaining the native conformation of antigens in liquid samples. Therefore, it is essential to validate antibodies for diagnostics in the specific assay format, under relevant conditions, with the appropriate sample type, and using the desired sample preparation protocol.

How does BioGenes ensure the development of relevant antibodies?

At BioGenes, we understand that the success of hybridoma production projects hinges on effective antigen design. Our process begins with a meticulous preliminary evaluation, where our team analyzes your project's unique requirements, considering the format of your intended application. Factors such as antigen immunogenicity, biological relevance, size (e.g., peptide), buffer conditions, potential cross-reactants, and antigen design are carefully assessed. This thorough analysis enables us to tailor the best solution to align with your project goals.

Moreover, we offer in the early development phase the option to provide you with parental clone supernatants. This allows you and your team to test them in your specific samples and assay conditions. 

Can cryocultures be stored with BioGenes?

BioGenes provides cryopreservation services for relevant hybridomas, offering convenient storage options for clients.

Who retains ownership of the generated clones?

The client retains exclusive ownership of the generated clones, ensuring that the results of the project belong solely to them.


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