Custom Antibody Production Principle

Having trouble finding an antibody? You can have an antibody custom-made for you

How to make Custom Antibodies

Immunogen preparation, immunization, hybridoma generation, screening, and purification are all processes used to generate particular antibodies. Antibody production, in a more particular sense, refers to the process of making antibodies.

When an antibody against a specific antigen is required but is not easily available from a provider, custom antibody manufacturing might be used. A 90 percent pure sample of antigen is all that is required to develop and harvest a tailored antibody against that antigen. If an antigen sample is not available, new peptides or proteins can be produced and antibodies developed against them. Custom antibody production procedures can create both polyclonal and monoclonal antibodies.

Boster Bio has established itself as a top antibody manufacturer after years of process development. We provide custom antibody production in gram scale volumes at highly competitive pricing and with a quick turnaround time. The product is delivered to your lab in just a few weeks, from gene sequence to purified antibodies.

Types of Antibodies

Antibodies are made from lymphocytes, these antibodies might be divided into two categories: monoclonal and polyclonal. Each one plays a critical role in the immune system, diagnostic tests, and treatment.

Multiple epitopes, or various locations on an antigen, can be recognized by polyclonal antibodies. They come from a variety of B cell lineages. Monoclonal antibodies are generated from a single B-cell line and identify only one epitope on an antigen.

how to produce Polyclonal abs

Polyclonal antibodies (pAbs) are made by injecting a specific antigen into laboratory animals like rabbits and goats. The animal is repeatedly immunized to produce greater titers of antigen-specific antibodies. These polyclonal antibodies can be harvested and collected from the antiserum in a matter of weeks.
Polyclonal antibody manufacturing is simpler and more cost-effective than monoclonal antibody production. Furthermore, polyclonal antisera can be made in a shorter time (4-8 weeks) than monoclonal antibodies (mAbs), which require 3 to 6 months to make.

Various animals and birds, including rats, hamsters, guinea pigs, chickens, goats, sheep, and donkeys, are used for immunization in addition to mice and rabbits. Immunoglobulin isotypes, immunoglobulin gene organization, antibody diversification mechanism, and antibody diversification organ sites differ between vertebrate animals.

Polyclonal Stepwise Production

Antigen preparation

The preparation of protein or peptide antigens is the first step in the manufacturing process. It's critical to assure the target antigen's quality, especially when binding to a conformational epitope is sought. The purity of the antigen used determines the specificity of the polyclonal antibody produced. Immunodominant impurities (1% ) may occur (e.g., with numerous bacterial antigens) and result in antibodies that are more active against the impurity than against the antigen of interest. Antigen purification is a time-consuming and difficult task.
Animal immunisation

The amount of antiserum required, the evolutionary distance between the species from which the protein of interest was generated and the species of the animal to be immunized, and prior experience with the immunogens all influence the animals used to produce polyclonal antibodies. Rabbits are the most commonly used animal since they are genetically distinct from the human and mouse sources of the proteins investigated. Rabbits can bleed up to 25 cc of serum each bleed without experiencing any negative consequences.

To boost the immune response for pAb generation, adjuvants are utilized. Freund's Adjuvant is a widely utilized adjuvant in scientific studies. The protein antigen is injected intramuscularly, intradermally, or subcutaneously into an animal of the specified species in the presence of adjuvant. Booster vaccines are given 4–8 weeks following the priming immunization and are repeated every 2–3 weeks. The animal is bled and whole blood serum is prepared prior to the priming immunization, as well as after each primary and booster immunization. Polyclonal antibody production should be stopped after the antibody titer has reached an acceptable level.
Antibody purification

Purification using Protein A/G affinity can enhance Immunoglobulin G (IgG) in raw antiserum while removing the majority of undesirable proteins. However, these preparations still include a significant amount of non-specific IgGs. When the antibody is employed in assays like ELISA, immunohistochemistry, and Western blot, the background noise will be significantly increased.
Antigen-specific affinity purification is frequently used to separate particular polyclonal antibodies from antiserum. Antigen affinity purification eliminates the majority of the non-specific IgG fraction, enriching the fraction of immunoglobulin that responds specifically with the target antigen.
Quality Control

To assure the quality of polyclonal antibodies, a set of quality control tests are performed after purification. Absorption at 280 nm is used to determine antibody concentration (A280). SDS-PAGE is used to determine the purity of polyclonal antibodies. An ELISA test was the best way for determining the polyclonal antibody titer.

how to produce Monoclonal abs

Monoclonal antibodies are a collection of antibodies produced by a single B cell clone. Milstein and Köhler were the first to create monoclonal antibodies in 1975. Hybridoma technology is the name for this process of producing monoclonal antibodies. Monoclonal antibodies have been widely used in biomedical research and therapeutic applications since that time.

The use of phage display, which was discovered by G. Smith in 1985, is another way of producing monoclonal antibodies. It's also become one of the most efficient methods for mass-producing peptides, proteins, and antibodies.Despite the fact that monoclonal antibodies are created against the same antigen, each clone reacts to various epitopes on the same antigen. In addition, each clone has its own set of programs. When choosing a monoclonal antibody, it's crucial to remember the antigen and clone names.

monoclonal Stepwise Production

Animal Immunisation

In hybridoma technology, the initial step is to immunize an animal (typically a mouse) with the right antigen. The antigen is injected subcutaneously, coupled with an adjuvant such as Freund's complete or incomplete adjuvant (adjuvants are non-specific potentiators of specific immune responses). The injections are done several times at different locations.
This allows B-lymphocytes that are reacting to the antigen to be stimulated more effectively. A final dosage of antigen is given intravenously three days before the animal is killed. This method has allowed immune-stimulated cells to expand to their full potential for antibody production. In the assay, the concentration of the desired antibodies is determined.
Fusion & Selection

The lymphocytes are completely cleansed and combined with HGPRT-defective myeloma cells. Because polyethylene glycol (PEG) is poisonous, the mixture of cells is exposed to it for a brief time (a few minutes). Washing removes the PEG, and the cells are retained in a new medium. A mixture of hybridomas (fused cells), free myeloma cells, and free lymphocytes make up these cells.
Only the hybridoma cells thrive when the cells are cultivated in HAT media, while the others progressively fade away. This occurs after 7–10 days of cultivation. It's critical to choose a single antibody-producing hybrid cell. If the hybridomas are isolated and cultivated separately, this is achievable.
Antibody purification

Before monoclonal antibodies may be employed for a number of applications, they may need to be purified. Protein Monoclonal antibodies are purified using affinity chromatography. It's the gold standard for monoclonal antibody (mAb) purification, and it's a technology that's attracting a lot of attention due to its superior performance and capabilities. Further purification procedures can be used if consumers require a higher level of purity.
Quality Control

To ensure the quality of monoclonal antibodies, a set of quality control tests are performed after purification. Absorption at 280 nm is used to determine antibody concentration (A280). SDS-PAGE is used to determine the purity of monoclonal antibodies. An ELISA test was the best way for determining the monoclonal antibody titer.

Primary Antibodies

Known for their high specifity and affinity, hundreds of primary antibodies that have been tested for several applications are available from Boster

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Polyclonal Antibodies

Custom polyclonal antibody generation service starting at $800. Service options are fully customizable. Contact us today to get a free consultation .

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Monoclonal antibodies

antibody design and production will ensure you the best chance of getting a high affinity monoclonal antibody that works for your application.

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Custom Antibodies

Boster Bio offers a special $600 custom antibody production package for customers who need custom antibodies for COVID-19 research, using rare species OR antibodies that have been discontinued by other suppliers.

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