Service and fees

Service

We provide the full range of services starting from library production and binder selection to characterization of target-specific binders.

 

Our immunisation strategy includes 4 injections of antigen administered in a fortnight interval. To maximise chances of raising a good immune response against all injected targets, we practice immunisation with maximum of 3 purified antigens per animal. The success of raising a specific immune response depends greatly on the nature and the quality of the target immunogen and it is serologically monitored for the increasing titre of IgG2 and IgG3 subclass (heavy-chain-only antibody).

Animal ordinance requires that immunogens are free from toxic and irritating substances (DTT, urea, guanidine hydrochloride, sodium azide, etc.).

 

Following immunisation, approximately 60-80 ml blood is collected to obtain lymphocyte RNA. VHHs genetic material is then cloned into a phage display vector. Routinely, phage libraries of 1x106 - 1x108 individual colonies are obtained which are then screened in consecutive rounds of binding, elution and amplification of phages which specifically bind to the target. Usually 2-4 rounds of panning are necessary to sufficiently enrich the phages of interest. This phase requires one month time and is succeeded by ELISA screening of at least two microtiter plates (192 colonies) per target.

 

Subsequent characterization of variability of identified binders from the nanobody library is done by sequencing of ELISA-positive clones.

The nanobodies of interest can be expressed recombinantly using different expression systems and purified using affinity chromatography.

Nanobodies can be  characterized in analytical and functional assays of choice (western blot, ELISA, size-exclusion chromatography, immunoprecipitation, immunohistochemistry).