![]() Anything over 60% has a strong likelihood of cross-reacting, but will need to be verified in an assay and sample specific manner.ĭoes my antibody cross-react with other species? What is the “right number” for homology to predict cross-reactivity?īased on our extensive experience in antibody development and working with thousands of customers, the magic number for almost guaranteed cross-reactivity is 75% homology with the immunogen sequence. ![]() 95% of Proteintech's antibodies are raised against recombinant, long-length human proteins, meaning that they have an increased chance of cross-reactivity in non-human models such as mouse, rat, drosophila and zebrafish. Conversely, if you are looking for an antibody to work in other species, use a polyclonal antibody against a recombinant, long-length protein. Therefore if you suspect your antibody of cross-reacting negatively, consider switching to a monoclonal antibody (a homologous IgG antibody population only recognizing one epitope). Remember that polyclonal antibodies have a higher chance or cross-reactivity due to the fact that they recognise multiple epitopes along the immunogen sequence. ![]() Cross-reactivity is not always a negative quality, for example cross-reactivity of an antibody for a target across species allows the same antibody to be used in multiple model organisms. Here, you can check for an antibody's potential cross reactivity against that of similar proteins or in other species. This can be achieved by pair-wise sequence alignment using NCBI-BLAST. Thus, testing an antibody for cross-reactivity with closely related proteins is a to ensure accurate results.Ī quick and easy check for antibody cross reactivity is to assess the percentage homology of the antibody immunogen to that of other similar proteins. This is often the case when two antigens have similar structural regions that the antibody recognizes.Ĭross-reactivity can invalidate the results of an experiment and has negative impacts on scientific reproducibility. Cross-reactivity between antigens occurs when an antibody raised against one specific antigen has a competing high affinity toward a different antigen, therefore the antibody is able to recognize a protein which is different to the one it was raised against. So overall, in general terms, antibody specificity helps us discuss if the antibody that we are using in our research (whether it’s a single antibody from a B cell or a pool of antibodies from serum) is able to detect our target protein specifically without cross-reacting with non-specific proteins.An antibody has a specific amino acid sequence (the Fab region) that dictates its affinity for a specific antigen. Techniques for increasing specificity will be discussed on the Monoclonal Antibody Alternative page. These two antibody production techniques will be discussed on the Polyclonal vs Monoclonal Antibodies page.Īlso, because serum contains antibodies against so many antigens (not just the protein that may have been used for immunizations), specificity is typically very low with raw serum. This brings up the issue of how antibodies are typically used, since they can be collected directly from the serum or by isolating the B cell(s) that is producing the antibody of interest. So, one antibody could potentially recognize two or more proteins if these proteins are highly homologous and contain the same epitope.Īlso, it is important to recognize that multiple antibodies will be generated against a typical protein antigen and so any one of these antibodies could potentially cross-react with another protein that contains the same epitope(s). However, it is important to recognize that a particular epitope could potentially appear on more than one protein antigen. This specificity allows precise detection of a target antigen such as a protein while avoiding detection of unrelated proteins that are not of interest. Each individual antibody protein is capable of binding specifically with one unique epitope thanks to the unique Antigen Binding Site located at the tip of the variable region on the antibody.
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