Why is it Important to Incorporate an Isotype Control Treated Group in an in vivo Experiment?
Incorporating an isotype-control-treated group is required to generate reliable data because it allows the researcher to accurately differentiate between results observed from primary antibody binding in an antigen-specific manner and results observed from non-antigen-specific binding or other nonspecific effects of antibody injection.
Primary monoclonal antibodies administered to animals can interact non-antigen specifically with Fc receptors on cells, including B cells, dendritic cells, NK cells, macrophages, and more. Fc receptor binding is sometimes involved in the primary antibodies’ mechanism of action, such as antibody-directed cytotoxicity (ADCC). Nevertheless, non-antigen-specific Fc receptor engagement can result in observable biological effects or phenotypes unrelated to antigen-specific binding. An isotype control antibody matching the isotype and subclass of the primary antibody will also bind to Fc receptors, allowing the researcher to monitor for those effects.
In addition to matching the isotype and subclass, an ideal isotype control must also match the primary antibody’s host species. Many primary monoclonal antibodies used for in vivo research in mouse models are created using hybridoma technology in rats or hamsters. When a xenogeneic rat or hamster IgG is injected into a mouse, an immune response may develop against the injected antibody. Multiple injections increase this possibility. As with Fc receptor binding, this can also result in observable phenotypes unrelated to antigen-specific binding. Including a group of mice treated with an isotype control antibody matching the primary antibody’s host species allows the researcher to monitor for these non-antigen-specific effects.
Using Isotype Control Antibodies
Selecting and using the correct isotype control antibody is essential to all in vivo experiments. These antibodies are used as a negative control – they match the exact host species, isotype, and subclass of the primary targeting antibody. However, they have no specificity to proteins present in the target organism or species. Isotype control antibodies are designed to be non-binding in in vivo animal models but retain all the nonspecific characteristics of the primary antibodies used in an experiment.
How to Choose an Appropriate Isotype Control Antibody
The isotype control antibody must match the host species, isotype, and subclass of the primary antibody. For the convenience of our customers, we have included this essential product information and the recommended isotype control for all of our primary antibodies on each product page. The table below is a comprehensive list of Bio X Cell’s isotype control antibodies suitable for both in vivo and in vitro research.
Why Choose Bio X Cell Isotype Control Antibodies?
Bio X Cell provides an extensive range of isotype control antibodies formulated for both in vivo and in vitro applications. Our isotype controls feature:
Exceptional Purity
Our optimized proprietary antibody manufacturing method ensures an ultra-pure antibody solution without added proteins or chemicals. Each lot is QC tested for purity using SDS-PAGE.
Ultra-low Endotoxin Levels
The level of endotoxin is QC tested for each lot. Our InVivoMAb™ products are ≤1EU/mg, and InVivoPlus™ products are ≤0.5EU/mg. Please contact our technical support for details if endotoxin levels below 0.5EU/mg are required.
Pathogen Free
Each lot of InVivoPlus™ product is screened for an exhaustive panel of murine pathogens. The results are detailed on product-specific datasheets to help you adhere to IACUC and Animal Facility requirements.
Low Protein Aggregation
Our proprietary antibody manufacturing method ensures an antibody solution with very low levels of protein aggregation. Additionally, each lot of InVivoPlus™ product is QC tested for aggregate level and guaranteed to be below 5% of the total protein.
Frequently Asked Questions About Isotype Controls
Using an untreated or PBS-treated group as a simplified negative control is not advised. An isotype control antibody must be used to accurately discriminate between results observed due to primary antibody binding in an antigen-specific manner and results observed due to non-antigen-specific binding or other nonspecific interactions.
Since the host species, isotype, and subclass of an antibody dictate its potential non-specific effects, the isotype control group must be treated identically to the primary antibody-treated group. For example, if the primary group is injected with rat IgG2a and rat IgG2b antibodies, then the isotype control group must also be injected with rat IgG2a and rat IgG2b antibodies.
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