• Phase I: Antigen Design and Immunisation (6–10 weeks)

APS offers detailed consultation with the customer in the design and preparation of protein or cellular antigens or advice with peptide synthesis and conjugation.

Immunisation schedules are determined using standard APS SOPs or customer SOPs where preferred.

Final stage determination and quantification of serum titre i.e. by ELISA, Western Blot, IP.  Samples may be supplied to customer for assay, if preferred.

• Phase II: Cell Fusion and Screening of Candidate Colonies (3–6 weeks)

Hybridoma fusion using standard APS SOPs or client SOPs where preferred into 10x96 well plates.

Initial screen by ELISA to select strongest positive candidate 'binders'. There is opportunity at this stage to select isotype specific antibodies

Expansion into 24-well plates for harvest of tissue culture supernatant for further antibody screening (ELISA, Western Blot or FACS)

At this point customers may opt to assess antibody performance and  5ml of supernantant from best 15–20 colonies may be supplied for this purpose.

• Phase III: Limiting Dilution Cloning and Cryogenic Preservation (3–5 weeks)

Limiting dilution cloning performed with strongest parental colonies as determined from Phase II.

Subclones with high specificity expanded and cryopreserved (standard preparation is 5 vials per subclone)

Shipment of hybridoma cells and up to 5ml of supernatant from each subcloned cell line

• Phase IV: Large Scale Antibody Production (3–6 weeks)

Milligramme to gramme quantities of anibody may be produced as required.  Production is performed in vitro using a variety of bioreactor systems with option for protein-free adaptation

Purification may also be carried out using protein A or G affinity chromatography