Optional VHH Services

QBI can optionally perform additional rounds of solid-phase panning beyond the initial three rounds typically scoped for VHH projects.  These rounds can enable further discovery and more fine-tuned discernment of high-affinity binders.  Binders that are better characterized before we go into the ELISA plate screening.

Working closely with you, we design a negative pre-panning step into our workflow to remove unwanted binders.  Negative pre-panning is recommended for removing phage-expressing VHHs that bind to parental cell lines or close sequences or species homologs that are not desired.  QBI will monitor enrichment using a phage titer dot assay to determine which round will be best to screen.

QBI can optionally add molecular tags to one or more of the high-binding VHH clones of Client’s choice and supply the tagged clones to Client.  Protein tags, such as his, avi, and myc, as well as common fluorescent stains, can be added in-house.  For non-protein chemical tags, such as dyes, we will subcontract with a third-party laboratory and bill Client at our cost. 

QBI can optionally generate humanized versions of the identified high-binding VHHs. We will use gene synthesis techniques to engineer the identified high-binder CDRs onto human VHH scaffolding. Service includes design, gene synthesis, and phagemid cloning of discovered VHH CDRs onto a humanized scaffold 

QBI can also optionally perform additional ELISA screening to verify that humanization has minimal/no effect on VHH binding affinity to targets.

Of course, we cannot make any guarantees about binder discoverability or affinity upfront.  Based on experience, our synthetic phage-display library of 10^9 clones yields Kd's in the double-digit nM range for typical protein targets.  If this is not sufficient for your needs, affinity maturation, via random mutagenesis using error-prone PCR performed on selected sequences, is an option.  We can generate "sub-libraries" that can yield around single-digit nM Kd's.

Additional strategies may be of interest to our clients.  If sequencing information offers insight into important CDR amino acids, site saturation mutagenesis is another option we can explore.  Alanine scanning is yet another, where we can "knock out" CDR residues with an inert amino acid.  Let us know if you wish to discuss these in further detail!

A benefit of our workflow approach is that you don’t have to commit to affinity maturation or any other value-added services at the start of the project.  If we run our standard workflow and produce binders that meet your needs, then we are done.  If they don't, then we can consider affinity maturation options and run that under a change order.