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HYBRIDOMA DEVELOPMENT SERVICES

Home / Antibody production / Hybridoma development

Thanks to a success rate of over 98% on more than 300 monoclonal antibody generation, ProteoGenix is able to offer the strongest guarantees on the market for hybridoma development. Sharing this success with you by proposing the charge only upon success in your application and in your hands makes us particularly proud!

Why choose ProteoGenix for your hybridoma development ?

Guaranteed hybridoma development
Strongest guarantees

You test purified antibodies and pay only if you are satisfied! Best guarantees on the market!

Successful hybridoma generation
High success rate

More than 300 monoclonal antibody generation and over 98% success rate!

Animal immunization for antibody generation
Immunization approach diversity

Protein, peptide, DNA immunization… We carry out all types of immunization strategies!

aaalac hybridoma production
AAALAC accreditation

ProteoGenix applies the highest ethical standards and is engaged for responsible animal use in science.

Application guaranteed antibody
Screening in target application

Developing a diagnostic tool? Flow cytometry, Sandwich ELISA, IHC, IF, IP, WB… Your antibody is guaranteed in the application of your choice!

Therapeutic antibody expert
Therapeutic antibody expert

Looking for therapeutic antibody development? Reformatting (bispecific, ADC), humanization, affinity maturation… Use our wide range of services and go straight to the clinic!

Choose your guaranteed hybridoma development package

Which application do you need a monoclonal antibody for? Click on the corresponding application below to discover the content and guarantees of our hybridoma development packages.

Therapeutic antibody development
Therapeutic antibody development
IHF – Immunohistochemistry IHC guaranteed antibody services
Western Blot - WB guaranteed antibody services
Modification specific guaranteed hybridoma services
Immunoprecipitation - IP guaranteed antibody services
Immunofluorescence - IF guaranteed antibody services
Flow cytometry - FC guaranteed antibody services
ELISA guaranteed antibody services
Sandwich ELISA guaranteed hybridoma services
Package name Code Gene synthesis & antigen production Immunization, fusion, ELISA screening & subcloning Screening in application Purified Ab production Antibody conjugation & ELISA pair identification
Standard ELISA ATX-PACK1M X
Premium ELISA guaranteed ATX-PACK2M X X X
Premium WB guaranteed ATX-PACK3M X X X X
Premium Flow Cytometry guaranteed ATX-PACK4M X X X X
Premium ELISA Sandwich guaranteed ATX-PACK5M X X X X X
Premium IF guaranteed ATX-PACK6M X X X X
Premium IP guaranteed ATX-PACK7M X X X X
Peptide guaranteed ATX-PACK8M X X X
Modification specific guaranteed package ATX-PACK9M X X X
Premium IHC guaranteed ATX-PACK10M X X X

Hybridoma development testimonial

"We collaborated on a project on generating a monoclonal antibody that recognizes a one-amino-acid mutation in a cancer-associated cell-surface protein. Although the project was technically challenging, we succeeded in generating a hybridoma clone that produced the desired antibody. In the course of the project, we found additional clones that recognized adjacent epitopes and ProteoGenix put forth an effort to perform additional subclonings in order to ensure the delivery of clones that perfectly work. In summary, ProteoGenix always strived to ensure maximal quality of the product and customer satisfaction, and the account Manager provided prompt and competent support at any time.”

Satisfied monoclonal antibody customer

Dr. rer. nat. Rudolf Übelhart, Project Leader, Ulm Institute of Immunology

Our hybridoma development service content

1
Antigen design for hybridoma generation
  • Antigen design

Definition of the most relevant immunization strategy.
Antigen design: peptide synthesis, gene synthesis & protein production in 2 systems or DNA Immunization.

2
Immunization fro hybridoma development
  • Immunization

Immunization of 5 mice with our optimized proprietary protocol

3
cell fusion for hybridoma generation
  • Cell fusion

Collection of the splenocytes from 2 mice for 2 fusions with a myeloma cell line

4
Hybridoma selection
  • Hybridoma selection and screening (polyclonal stage)

Hybrid cells selection (HAT selection)
Culture supernatant screening vs. target antigen (ELISA screening)

5
Hybridoma cells expansion and selection
  • Isolation and selection of the best monoclones

Isolation of monoclones by limiting dilution.
Expansion and screening of the monoclones by ELISA or in target application.

Case study: monoclonal antibody production for the detection of a human cell membrane protein

AIM OF THE PROJECT

A customer requested the production of a monoclonal antibody for the detection of a human cell membrane protein. Design of the antigen was particularly challenging as the protein was known to be difficult to express in mammalian cells.

Application guaranteed: Flow cytometry

 

ANTIGEN DESIGN AND PRODUCTION

  • A protein fragment estimated as the possible extra-cellular domain and possible to express in mammalian cells was designed.

  • The corresponding gene was designed and optimized for mammalian cell expression.

  • Mammalian cells were transfected with a plasmid expressing the target protein coupled to GFP.

Protein expression was performed both in HEK293 and in CHO cell lines. Final production was performed in HEK.

Production method: protein secreted in culture supernatant
Yield: 0.43mg/L
Quantity produced: 0.86mg
A final QC was performed by SDS-PAGE.

ANIMAL IMMUNIZATION AND SCREENING

5 mice were immunized with 4 injections containing the protein antigen. More information about the protocol are given in the complete report.

FC results of the mice serum tested against the immunogen:

Sample tested Positive control Mouse 3 Mouse 4
Mice serum after 3rd injection 90% 42% 29%
Mice serum after 4th injection 76% 47%

Results for the other mice are provided in the completed report.
Mouse 3 demonstrated the best immunogenic reactivity against the antigen in FC.
As our project includes 2 fusions, both mice 3 and 4 were selected to perform fusion.

SUPERNATANT SAMPLES AFTER FUSION (MOUSE 4): TEST IN ELISA

Confirming screening after fusion
Clone ID 65 66 67 68 69 70 71
OD value 1.254 1.615 1.306 1.412 1.224 1.198 1.334
Clone ID 72 73 74 75 76 77 78
OD value 1.251 1.011 1.394 1.225 1.636 0.153 1.647
Clone ID 79 80 81 82 83 84 85
OD value 1.07 1.223 1.267 1.847 1.395 1.282 1.783
Clone ID 86 87 88 89 Negative Positive
OD value 1.036 0.983 1.173 1.116 0.088 1.731

24 positive clones were selected in ELISA.
Determination of the best clones for FC is done directly in the target application.

 

SUPERNATANT SAMPLES AFTER FUSION (MOUSE 4): TEST IN FC

Details about the protocol are provided in the complete report.

Clone ID 65 66 67 68 69 70 71
FC result / % 37.48 1.27 20.27 62.50 1.18 11.40 1.38
Clone ID 72 73 74 75 76 77 78
FC result / % 75.09 1.01 6.50 56.76 73.16 1.18 1.09
Clone ID 79 80 81 82 83 84 85
FC result / % 1.07 1.05 1.49 6.75 71.92 1.18 1.02
Clone ID 86 87 88 Positive
FC result / % 1.04 1.01 7.36 80.73

11 positive clones were obtained after fusion in FC. These clones were subcloned for further antibody production.

Similar ELISA and FC analysis were performed on mouse 3. 14 positive clones were obtained after FC. Results are available in the complete report.

 

ANTIBODY PRODUCTION

30 clones were selected by our customer for further production. Supernatant samples were tested in ELISA and FC after 2 subcloning steps. Results are provided in the complete report.

 

CONCLUSION

  • We succeeded in producing a difficult-to-express protein in sufficient quantities to develop monoclonal antibodies. Several expression tests with several mammalian cells strains were performed to reach this goal.

  • The immunization protocol in mice allowed us to develop good antibodies against the antigen.

  • After 2 fusions, 30 hybridomas/clones detecting the antigen in flow cytometry were produced.

  • 30 clones and 30 purified antibodies were delivered to our customer.

     

Please click on the button below to get the complete report.

 

You would like to know more about our hybridoma generation services? Please check another anti-peptide case study.

What are the advantages of hybridoma development for antibody generation?

Hybridoma technology revolutionized the whole biotechnological field by providing a method allowing unlimited and reproducible monoclonal antibody production. Today, hybridoma development is still a reference method for the high sensitivity antibody generation. Therefore, hybridoma technology remains the best technique for assay development. It represents also a good alternative to phage display for therapeutic antibody development. However, administration of monoclonal antibodies produced by hybridoma technology lead to severe side effects due to their mouse origin. Thanks to recent advances in antibody engineering, the immunogenic mouse components can be reduced by a so-called antibody humanization process. Coupled to further affinity maturation, this global process can be highly relevant to generate optimized humanized antibodies suitable for therapeutic applications.