Isotype controls are a kind of antibodies that share the same isotype with the primary antibody but without specificity to the target antigen.
Researchers use they as negative controls to differentiate non-specific background signal from specific antibody signal. They are mainly used in flow cytometry or less in immunohistochemistry. In addition to this, whole molecule immunoglobulins can also be used as a control, working as a standard, coating protein or blocking agent in different assays, such as ELISA, immunoblotting, immunoprecipitation, etc.
However, people are saying that isotype controls might be replaced by unstained samples. Is it true? Theoretically it is accessible. But put it into practice, it is risky. For unstained samples, one might get ideal background signal for specific makers in some time and see some rise which makes you think all of them are specific. But how about the increase of the antibody receptors in the cell? In addition to this, there is other saying prevents you from using non-specific staining and not having isotype controls. But once done experiments with isoptypes you will find the difference.
Although isotype control antibodies are not ideal for every experiment, they are still very useful and even necessary in many conditions. The following are examples of such situations:
- Help to properly adjust the cytometer.
- Help to evaluate unusual or unexpected staining patterns in specific antibody samples.
- Help to assess the up- or down-regulation of a particular surface marker after an in-vitro manipulation.
- Help to provide more and valuable information for samples with poor viability and contain large amounts of cellular debris.
- In flow cytometry experiments.
- In immunohistochemistry experiments.
“But it’s true that controls can be costly and annoying in terms of its importance of contribution to valid and reproducible experiments. To choose repeat an experiment or to do it right the first time is the choice.” Creative Diagnostics scientists said.
About Creative Diagnostics
Creative Diagnostics is a leading manufacturer and supplier of antibodies, viral antigens, innovative diagnostic components and critical assay reagents. Many isotype control products are available to be chosen at CD, such as purified antibodies from the same isotype and species as the primary antibody.