llama antibody

What Does It Take to Bring New Nano Antibodies (nAbs) to the Hands of Researchers?

Judging from the hundreds of papers published using camelid VHH antibodies as reagents, there are probably thousands of researchers who have experience with this type of antibodies by now. We like to call the ~15kD camelid VHH antibody nano antibody or nAbTM. Once someone experiences how well a nAb works for co-IP using a fluorescent protein as tag, they often wonder what it takes to bring nAbs to broader use.

The success of a nAb project starts with the antigen presentation. It is critical to build the capability to produce large quantities of recombinant antigen for immunization. At Allele, our scientists also established some unique presentation formats for traditionally difficult targets (e.g. large membrane proteins).

After llama immunization, the next step is screening. With the goal of creating large scale nano antibodies against diverse targets, we have developed multiple high throughput screening methods to cover very large, diverse libraries generated from immunized animals. The technologies will continue to evolve as the scale of nAb generation continues to expand. We have the ability to functionally screen for site-blocking antibodies and antibodies that only recognized natively folded targets, or targets in their naturally occurring presentations.

A nAb isolation project does not end with the obtaining of a cDNA clone. Or, if it does, the nAb is probably not as great as what Allele Biotech has been offering. In our hands, all nAbs go through an engineering step beginning with the generation of a 3D structural model of the isolated clone. We use structure-guided design to alter the protein, allowing us to improve its properties. This includes increasing affinity, solubility, or altering the protein to improve performance for specific applications. We also like to use known structures of traditional monoclonal antibodies to assist camelid VHH antibody engineering against specific targets.

With a finalized clone in hand, the next step is to establish protocols for commercial production. The Allele team spends a tremendous amount of effort aimed solely at high-yield, low-cost recombinant VHH antibody production in a variety of formats, so that the costs for other scientists to take advantage of these great reagents can be kept as low as possible.

Last but not the least, nAb labeling, including conjugating stable soluble VHH antibody to solid supports for immunoprecipitation or to fluorophores for detection, requires additional expertise and tight operation control. However, our vision is to have a modular system for antibody labeling that will enable the end user to select from a variety of fluorophores and other detection tags, which can be instantaneously and irreversibly coupled via simple mixing.

Note added: we work with commercial (diagnostic and clinical) partners from developing nAbs all the way to the market. We have expert scientists available to customers and licensees for consultation and troubleshooting antibody- and imaging-related questions and problems.

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Camelid Antibodies

When it was discovered that animals in the camel family produce antibodies with no light chains, the idea that a single-domain fragment can bind as well as a full 4-chain antibody formed a breakthrough. So far it has been a relatively less known one.

Smaller antibody fragments have been tested for therapeutic uses because classical IgG antibodies are too bulky to penetrate tissues well, and very expensive to produce. Different combinations of antigen-binding variable regions are used, e.g. scFv, Fab, diabody, all to some degree of success. In comparison, the N-terminal domain of camelid antibodies, termed VHH domain (nanobody, VHH antibody), represents a naturally evolved, only 13-15 kD in size, fully functional target binding fragment with many advantages.

The only other known species outside camelidae family that has heavy chain antibodies is particular cartilaginous fish, nurse shark. Although the arrangement of CDRs is somewhat different between the camel and shark heavy chain variable regions, they share many characteristics such as extremely high stability (maintaining functions after100 C heat and extreme pH treatment).

Accumulating reports have demonstrated the therapeutic potentials of camelid antibody-based fragments in treating cancer, neural diseases, even use in hair dandruff preventing shampoo. For basic research, the tiny antigen binders can be used as tools for quantitative pull down with unmatched efficiency, recognizing previously inaccessible enzyme cleft as antigens, and providing libraries for binding partner selection.

Allele Biotech has been working on display antibody selection from its early days through an NIH grant, and recently carried out an NIH/NCI contract for scFv yeast display.

Check out Allele’s current Camelid antibody products: http://www.allelebiotech.com/allele3/CM.php

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