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mAb - Hybridoma’s development

mAb- Hybridoma’s development

Eurogentec develops murine hybridomas in 16-19 weeks following a stepwise approach with testing sample phases. A reporting is sent after each phase and discussed with the customer for potential adaptation. If no positive hybridoma is obtained after phase 3, only phase 1 will be charged.

However, a personalised procedure may be designed and adapted to your needs.

mAb – Hybridoma’s generation Programme Overview

Programme Details

Project Development:

Prior to project initiation, you can benefit from our personalised project proposals and expertise.

Starting Material:

You send us:

  • Your purified protein
  • Or your peptide sequence
  • Or your protein’s accession number
  • Or other antigen

Phase I: Immunisation ~ 6 weeks

  • A minimum of four mice (6-8 weeks old) are used for immunisation.
  • Preimmune serum is taken on day 0.
  • The first injection on day 0 with equal parts (v/v) of complete Freund's adjuvant and antigen.
    The following boosts are carried out with equal parts (v/v) incomplete Freund's adjuvant and antigen on days 21 and 36 with an additional final boost prior to fusion.
  • You receive an ELISA report against the supplied antigen and serum sample.
  • Selection of the best responding host on the basis of Customer's test results.

Phase II: Fusion/Hybridoma production~ 2-3 weeks

The splenic lymphocytes of the best responding animal will be fused with the cell line Sp2/O-Ag-14 using PEG (polyethylene glycol), and the resulting hybridomas will be seeded in eight 96-well plates in HAT medium.

  • You receive a Fusion report.

Phase III: Screening for positive hybridomas ~ 4-5 weeks

  • Screening for positive antibody producing hybridomas (ELISA against the target antigen)
  • Positive colonies are expanded and tested for antibody production
    On request, positive colonies can also be screened against other antigens for cross-reactivity or against a specific region of the antigen of interest Different screening techniques are available and can be used on request. Please contact us for more information
  • You receive a screening report, an ELISA against your antigen, a delivery of >1 ml positive supernatants (Maximum 30) for screening confirmation and additional tests.

Phase IV: Cloning and isotyping of positive hybridomas ~ 4-5 weeks

  • The specific hybridomas selected by the customer at the end of the Phase III are cloned by serial dilution.
  • Positive hybridomas are collected in vials
  • Monoclonal antibody isotype is determined
  • You receive hybridomas ELISA and isotyping report, > 10 ml of supernatant from 1-2 positive clones and frozen hybridomas are delivered.

How to start

To qualify and build your project or to receive a quotation adapted to your needs, please contact your local representative or send us an e-mail at

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