What is an Anti-Idiotypic Antibody?

As shown in figure 1, an anti-idiotypic (Anti-ID) antibody binds to the idiotype of another antibody, usually an antibody drug. An idiotype can be defined as the specific combination of idiotopes present within an antibody's complement determining regions (CDRs). A single idiotope, is a specific region within an antibody's Fv region which binds to the paratope (antigenic epitope binding site) of a different antibody. Therefore, an idiotope can be considered almost synonymous with an antigenic determinant of an antibody.

Antibody idiotype and paratope interaction

A: Antibody Drug Targeting Antigenic Epitope

B: Anti-Idiotype Antibody targeting Antibody Drug

An idiotope within the CDR region of an antibody drug interacts with the paratope of an anti-idiotype antibody.

Figure 1. Structure of anti-idiotype antibody. (A) The paratope of an antibody drug lies within its CDR region and will bind to the epitope of a target antigen. (B) The paratope of an anti-ID will bind to the idiotope of its target antibody drug. The idiotope is within the antibody drug's CDR region and could be next to, identical to, or far from the antigen binding site.

Types of Anti-Idiotype Antibodies

There are three main types of anti-ID antibodies based on the way they interact and detect an antibody drug, as shown in figure 2. The first is an antigen blocking anti-ID antibody, named so because the antibody drug's paratope and idiotope overlap with one another. Because of this, the antibody drug's target antigen and the anti-ID antibody will compete with one another. Therefore, this form of anti-ID antibody will only detect free antibody drug. The second type of anti-ID antibody is called non blocking because the antibody drug's paratope and idiotope do not overlap. Therefore, the anti-ID antibody and the antigen can simultaneously bind to the antibody drug without affecting one another's binding capability. Because of this, non-blocking anti-ID antibodies are used to detect all forms of available antibody drug (free and antigen bound). The third type of anti-ID antibody is a complex specific anti-ID. This is because the anti-ID antibody cannot bind to the antibody drug unless the drug is already bound to its antigen. Therefore, this type of anti-ID antibody is only able to detect bound antibody drug.

Types of anti-idiotype antibodies

A: Antigen Blocking Anti-ID Antibody

B: Non-Blocking Anti-ID Antibody

C: Complex Specific Anti-ID Antibody

Different types of anti-idiotype antibodies may bind to free, bound or all available antibody drugs

Detects Free Drug

Not Paratope-Specific
Not Inhibitory
Detects Total Drug (free, partially bound, and fully bound)

Drug-Target Complex Specific
Not Inhibitory
Detects Bound Drug

Figure 2. Types of anti-idiotype antibodies. (A) Antigen blocking anti-IDs bind directly to the antigen binding site of an antibody drug, directly competing with target antigen. (B) The idiotype which non-blocking anti-IDs bind to does not overlap at all with the paratope of the antibody drug, allowing the antibody drug to bind its target antigen and an anti-ID simultaneously. (c) Complex specific anti-IDs will only bind to the idiotope of an antibody drug when that drug is already bound to its appropriate antigen.

Uses of Anti-Idiotype Antibodies

Pharmacokinetic Assays

Since most anti-ID antibodies are generated against a specific antibody drug, they are commonly used in preclinical practices for pharmacokinetic (PK) analysis. PK is the study of drug metabolism throughout the body. Specifically, clinicians will determine the rate of drug absorption, distribution, bioavailability, and excretion in various cohorts of patients. In order to accomplish this, researchers need to be able to track antibody drugs which are bound or unbound to their designated target at various time points post delivery. Through the use of anti-IDs, primarily mAbs, various forms of antibody therapeutics can be easily tracked and quantified in patient serum, blood, urine, or other bodily fluids. Examples of pharmacokinetic assays (ELISA format) using anti-IDs are shown in figure 3.

Antibody drug ELISA based PK assays

Four commonly used ELISA based PK assays use anti-ID antibodies to detect antibody drugs in human serum

Figure 3. Types of ELISA Based PK Assays. There are four commonly used formats of ELISA based PK assays. (1) The antigen capture assay involves an antigen coated plate to capture antibody drug (e.g., from human serum). A labeled anti-Fc is then washed over to bind the antibody drug and visualized. (2) An anti-ID bridging assay involves an anti-ID coated plate that binds the antibody drug and is washed over with a labeled anti-ID for detection. (3) An anti-ID capture sandwich assay involves an anti-ID coated plate that binds the antibody drug and is washed over with a labeled anti-constant region IgG. (4) An anti-ID antigen-bridging assay involves an anti-ID coated plate used to capture the antibody drug bound to a labeled antigen.

Immunogenicity Assays

Polyclonal anti-IDs are commonly used for immunogenicity assays as reference controls. Immunogenicity is the ability of a therapeutic, such as an antibody drug, to induce a humoral and/or cell-mediated immune response, which leads to development of anti-drug- antibodies (ADAs). Anti-drug antibodies lead to the negation of all antibody drug related effects, essentially completely inhibiting the therapeutic aspect of the drug. Therefore, it is extremely important for researchers to analyze the immunogenicity of a new antibody drug during preclinical analysis. An anti-drug antibody is very similar to an anti-ID since both bind to the same antibody drug, therefore a pool of polyclonal anti-IDs can be used as positive controls when analyzing for the presence of anti-drug antibodies within patient samples. An example of an ELISA based immunogenicity assay using an anti-ID as a positive control is shown in figure 4.

Antibody drug ELISA based PK assays

ELISA based immunogenicity assays detect anti-drug antibodies that may arise following use of antibody drugs

Figure 4. Types of Immunogenicity Assays. (A) If an antibody drug induces the development of anti-drug antibodies, they can be detected through a direct binding ELISA. In this assay, an antibody drug coated plate is washed with serum. The plate is then washed with a labeled IgG, which only binds to the constant region of anti-drug antibodies. Therefore, the ELISA will only fluoresce if there are anti-drug antibodies present in the sample serum. (B) In a bridging ELISA assay, an antibody drug coated plate is exposed to serum allowing anti-drug antibody capture. The same antibody drug, but labeled, is used for detection of the anti-drug antibodies through binding to their ScFv region. Both of these assays need to be analyzed via a positive control, such as anti-IDs.

GenScripts Anti-Idiotype Antibody Generation Packages

GenScript now offers distinct anti-ID generation packages, which are specific to the downstream application, host species, and clonality of your choosing. GenScript's anti-ID generation packages include:

Rodent mAb: Immunize mice or rats with your antibody drug to develop strong anti-ID antibodies generated in as little as 3 months time for the perfect price! The deliverables include stable hybridoma cell lines, purified mAb, and a fully comprehensive COA report analyzing blocking type and specificity testing.

Rabbit mAb: Anti-ID antibodies generated using GenScripts MonoRab™ platform bind with the highest specificity and sensitivity on the market. After in house parental clone supernatant analysis, positive clones go through V region sequencing, allowing for optimum recombinant expression of your selected mAb. This package also includes guaranteed delivery of a second biotin conjugated anti-ID mAb pair to allow for easy generation of sandwich ELISA based PK analysis.

Rabbit pAb: GenScript's rabbit pAbs provide a convenient turn around time and price range for customers interested in using polyclonal anti-IDs for immunogenicity analysis. Using our express antibody generation protocol and extensive purification and screening process, GenScript rabbit anti-ID pAbs are a reliable option for your next project.

Click here to learn about ready-made anti-idiotype antibody products against blockbuster drugs.

Anti-idiotype Antibody Services

√ Guaranteed application in immunoassay development for PK and ADA analysis
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