Llama Monoclonal Antibodies
Camels and their South American close relatives, llamas and alpacas (Camelidae) have antibodies that are unusually small and can penetrate tissues that larger antibodies cannot. Llamas antibodies are thus offering new insights into many different aspects of research.
But why llamas?
Llama antibodies have all the characteristics we expect from mammalian immune systems. However, they also include a group of smaller antibodies that don't correspond to anything seen in other animals. Raymond Hamers and Cecil Casterman found that these antibodies were not degraded forms of larger antibodies, but were unique and functional. They are heavy chain dimers with an extensive antigen-binding repertoire. They lack the light chains altogether. Because of their small size, single chain antibodies raised against small enzymes such as lysozymes or ribonucleases can access their enzymatic active sites to provide potent enzyme inhibition.
In conventional antibodies, antigen recognition is provided by both heavy and light chains. The smaller single chain antibodies bond in narrow regions of a molecule, allowing it to reach parts of a molecule that other antibodies can't. The binding domain (V-region) of camel or llama antibodies can be effectively engineered to a unique, small single domain antibody, sdAb or nanobody. These small antibodies can more easily enter a localized tumor. What we have is a different tool, that can be put to use in specialized applications.
With our novel Single Plasma Cell Interrogation (SPIN®) Technology, single chain antibodies are isolated from individual plasma cells of immunized llamas and are expressed in mammalian cells, allowing them to be functionally analyzed. This technique leads to broad epitope coverage and high-affinity antibodies allowing fast, efficient and economical development of these unique antibodies.
Advantages of SPIN® Technology
1. High affinity: Isolating Plasma cells, the most mature and terminal differentiated B cells for library construction ensure high antibody affinity.
2. Straight forward: Direct expression in mammalian cells and screening of antibody individually allows for superior results and analysis.
3. Broad epitope converage: Assays are made functional by screening targets directly using indirect ELISAs.
Llama IgG Gene Structure
The Arabian camel (Camelus dromedarius) and the llama (Lama glama) have three IgG subclasses: the conventional IgG1 and the non-conventional IgG2 and IgG3.
What makes IgG2 and IgG3 different from its two counterpart? There's a splicing defect in the CH1 exon which causes the CH1 domain to be absent in both the gamma 2 and gamma 3 chains. This leads to a striking difference from IgG1: a lack of light chains in IgG2 and IgG3.
Advantages of llama VHH single domain antibodies:
1. Accessibility: VHH single domain antibodies are only 15kD in size, compared to about 150kD for typical immunoglobulins. The smaller size makes it easier to penetrate tissues and gives them access to “hidden” epitopes while readily crossing the blood-brain barrier.
2. Stability: VHH antibodies are extremely stable and resistant to both high acidity and high temperature. They're even able to fold back into a functional protein after denaturation.
3. Solubility: Single domain antibodies display a marked reduction of in hydrophobicity, which in turn leads to a very high solubility.
4. Engineerable: VHH antibodies are pure monomers, offering fast-track lead optimization and engineering to bi-specific nanobodies.
|Phase I||Immunization||Immunization of one Llama on standard protocol ELISA to assess the titer|
|Phase II||Plasma Cell Isolation||SPIN® technology isolates plasma cells. ONLY the terminal differentiated plasma cells with the most mature antibodies will be applied for library construction|
|Phase III||Single Domain Library Construction||Single domain antibody genes are amplified to construct small libraries|
|Phase IV||Expression and Screening||Directly screening individual single domain antibodies using ELISA or functional assays make antibodies suitable for downstream work.|
|Phase V||Production||Efficient HEK293 or CHO cell expression|
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