Immunisation and Hybridoma Preparation
Immunisation can begin immediately once the immunogen has been successfully designed and produced. The schedule below gives an idea of a minimum time scale for the production of a monoclonal antibody.
1st immunisation
Day 1
2nd immunisation
Day 11
3rd immunisation
Day 21
Test bleed to check serum reactivity
Day 28
Final booster immunisation
Day 31 - 33
Spleen cells harvested and cell fusion performed
Day 33 - 36
Primary screening of hybridoma colonies, usually by ELISA
Day 40 – 54
End of primary screening and identification of potential antibodies of interest
Day 50 - 54
Expansion of individual hybridoma colonies producing relevant antibodies and re-screening for reactivity with the target antigen
Day 40 – 54
The ELISA positive supernatants are further characterised by testing against mammalian cell transfectants that express the target protein. This helps to ensure antibody specificity
Day 40 - 54
Cloning of hybridoma cell lines secreting specific monoclonal antibodies and storage in liquid nitrogen
Day 45 - 60
Culture of clone(s) to produce large volumes of antibody (in the form of supernatant) for further characterisation
Day 60 -75
Further characterisation available on request: Western blotting and IHC studies, etc.