Over the past few decades, the field of antibody research and application has undergone revolutionary changes. In this process, a special source of antibodies - single-chain antibodies (also known as nanobodies) from camelids (including alpacas) have attracted widespread attention from scientists around the world.

The immune system of camelids such as alpacas is unique. In addition to conventional heavy and light chains, they can also produce antibodies with only heavy chains. These heavy-chain antibodies are similar to conventional antibodies in function, but have the advantages of small size, high temperature resistance and stability, and strong tissue penetration ability. It is precisely because of these characteristics that nanobodies have great potential in biomedicine, scientific research, diagnosis and many other fields. With the continuous deepening of biomedical research, nanobodies have become a hot spot in the market due to their unique biophysical properties and wide application potential.

一. Reasons and Advantages for Choosing Alpaca Immunization:

1. Biophysical properties: Alpaca nanoantibodies are small in size and can reach deep into cells and tissues that are difficult for traditional antibodies to reach. Alpaca nanoantibodies show excellent stability at extreme temperatures and pH values, are highly water-soluble, and are suitable for a variety of application scenarios, making them an ideal choice for many applications that traditional antibodies cannot handle.

2. Easy to engineer and produce: The simple structure of nanoantibodies makes it easier to engineer, including humanization, multivalent construction, etc., and can be efficiently expressed in bacteria or yeast systems to reduce production costs.

3. High specificity and high affinity: The immune system of alpacas can produce antibodies with high specificity and affinity for specific antigens, which is crucial for the development of therapies and diagnostics.

二. Alpaca Immunization Process:

1. Pre-screening and Preparation:

①. Animal selection: Select alpacas of appropriate age and physical health from a healthy group, and perform blood tests and disease screening on them to ensure that there are no infectious diseases or other health problems.

②. Preparation of immunogens: Immunogens are the key to inducing immune responses. Appropriate proteins, peptides or other suitable substances need to be selected according to the needs of target antibodies.

2. Immunization, Blood Collection and Serum Separation:

①. Immunize alpacas according to the immunogens we prepared and the immunization flow chart. Each immunization is generally fourteen days apart.

②. Collect negative serum before immunization, and collect blood one week after four or five immunizations for immune titer detection (the collected blood is centrifuged for 10 minutes, and the upper serum is separated and preserved). Specific antibodies and titers are detected by ELISA, Western blotting or other immunoassay methods.

3. Fusion and Cloning (if monoclonal antibodies are required):

①. Obtain B lymphocytes from alpacas and fuse them with myeloma cells to produce hybridoma cells.

②. Screen hybridoma cells to find clones that produce the required specific antibodies.

4. Antibody Production and Purification:

Produce antibodies by culturing hybridoma cells in vitro or using recombinant DNA technology in microorganisms, mammalian cells or other systems. Purify antibodies using protein A/G columns or specific affinity chromatography.

5. Quality Control: Perform quality checks on the final product, including tests for purity, specificity, affinity, and biological activity.

6. Product Delivery: Provide the required documents and product specifications and deliver the purified antibodies to the customer.

In addition, the immunization scheme, adjuvant selection, immunization schedule, immunogen dose, etc. need to be customized according to specific project requirements. After receiving the customer's immune antigen and passing QC verification, TechBio delivers the designed immunization plan to the customer, including blood collection plan, serum titer ELISA test plan, animal immunization dose, blood collection and PBMC separation plan, etc.

三. Requirements for Various Immunogens:

Proteins and peptides: require high purity, correct conformation, and sufficient immunogenicity to induce a strong immune response.

Small molecules: usually need to be coupled to a carrier protein to increase immunogenicity, and the coupling site is clearly indicated to correctly identify specific epitopes.

Complex samples (such as cells, viruses): require adequate characterization to ensure their integrity and activity, as well as safety assessment.

四. How to Obtain High-titer Serum and Subsequent Screening to Obtain High-affinity Antibodies:

1. High-quality Immunogen Preparation:

①. The purity of the immunogen should be as high as possible to avoid nonspecific reactions.

②. For protein immunogens, ensure that it remains intact in three-dimensional structure to better simulate the antigenic epitope in the natural state.

③. If the target is a surface antigen or receptor, consider using full-length, complex or recombinant protein expressed in an appropriate expression system.

2. Selection and Use of Adjuvants:

①. Use adjuvants that can enhance long-term immune memory and stimulate the production of higher antibody titers, such as Freund's complete or incomplete adjuvant, hydrated aluminum hydroxide, etc.

②. In some cases, in order to further enhance the immune response, specific immunostimulatory complexes (such as CpG oligonucleotides) can be added to the adjuvant.

3. Design of Immunization Plan:

①. Develop a reasonable immunization schedule, including basic immunization, multiple booster immunizations, and give enough time after each immunization to allow the immune system to fully respond.

②. Take blood samples at appropriate times to monitor antibody levels to determine whether additional booster immunization or adjustment of the immunization regimen is needed.

4. Serum Screening and Antibody Affinity Maturation:

①. Use ELISA, Western blotting or immunoprecipitation to screen high-titer serum samples.

②. For situations where monoclonal antibodies need to be generated, B cells can be isolated from immunized alpacas, single-cell screening can be performed, and then specific antibody genes can be cloned using molecular biological methods (such as PCR, plasmid construction, etc.).

③. If necessary, the affinity of the resulting antibodies can be enhanced by in vitro affinity maturation technology (for example, based on phage or yeast display technology).

5. Production and Verification of High-affinity Antibodies:

①. Use appropriate expression systems (such as mammalian cells, Escherichia coli, etc.) to produce recombinant antibodies.

②. Purify antibodies and conduct comprehensive biological function and binding property analysis to ensure that they have the expected high affinity and specificity.

With its deep expertise and resources in nano-antibody research and development, Tek Biotech is committed to providing customers with high-quality services, especially in the key technology of alpaca immunization. Our self-developed adjuvant is carefully optimized for the immune system of alpacas to ensure that it has a potency and specificity that exceeds traditional adjuvants. With this unique adjuvant, we promise to complete the immunization program within the specified time and deliver high-titer serum. And every project, no matter how big or small, will benefit from our strict quality control process to ensure that every antibody delivered to customers meets the highest standards. In addition, Tek Biotech has a large group of healthy alpacas to provide strong support for a variety of immunization projects, and we always adhere to the principles of animal health and welfare. Alpaca immunization technology occupies a core position in the discovery and development of nano-antibodies. With this technology, Tek Biotech creates high-quality nano-antibody screening and production services for customers, helping to smoothly transition scientific research results from the laboratory to clinical applications to meet market needs.