An antibody to biotin is not bred against biotin, but is an antibody conjugated to biotin, which binds to avidin for use in scientific studies. Biotin can be conjugated to monoclonal or polyclonal antibodies and used to detect antigens through fluorescent imaging. Biotin can also be used to detect neuronal receptors affected by neurotoxins or to detect cellular mRNA regulation. Biotinylated antibodies can be purchased or produced through online protocols.
Contrary to what one might think at first glance, an antibody to biotin is not a substance bred against biotin. Rather, an antibody to biotin is an antibody that has been conjugated to biotin. Biotin binds strongly to a ligand – avidin – which can, in turn, be conjugated to light-detectable molecules such as fluorophores. Biotin and avidin form one of the strongest biologically covalent bonds known, making the ligand/receptor units conjugated to biotin and avidin particularly useful in scientific studies.
Biotin is a vitamin of the B complex, vitamin B7. It is such a small molecule that multiple biotin molecules can be conjugated to a single monoclonal or polyclonal antibody. The fact that biotin is also water soluble allows for more versatility in buffer systems than if it were insoluble. For greater specificity, monoclonal antibody conjugation to biotin is often recommended over polyclonal antibody conjugation. The biotin molecule can be directly bound to the antibody of interest, or it can be linked via a molecular tail to allow for greater freedom of movement.
The biotin ligand avidin is found in egg whites. It can be made cheaply and easily isolated. Multiple avidin molecules will bind to each biotin receptor, amplifying a single antibody sensing signal. When a biotin antibody adheres to an antigen of interest, an experimenter can then expose the complex to the fluorophore-conjugated avidin, making the complex detectable. Through fluorescent imaging, the biotin antibody and avidin-conjugated ligand can be detected by light microscopy, and the antigens of interest can be studied in situ.
While the creation and use of a biotin antibody and a fluorescently labeled avidin molecule is a common solution for antigen detection, there are alternatives that still make use of the biotin/avidin complex. Binding of biotin to a ligand such as a neurotoxin may be beneficial in detecting the location of neuronal receptors affected by the neurotoxin in question. It is also possible to conjugate biotin to a nucleotide sequence such as messenger ribonucleic acid (mRNA) to find out where these nucleotide sequences are translated into proteins or to detect cellular mRNA regulation.
It is possible to purchase biotinylated monoclonal or polyclonal antibodies, although the production of biotinylated antibodies is within the reach of many experimenters. Protocols can be found online suggesting methods of biotinylation of the antibody, although protocols more easily followed link biotin directly to an antibody rather than suggesting how to intercalate a molecular spacer. The molecular spacers between biotin and its associated antibody allow for greater distance to expose more molecules to binding sites, and these biotin antibodies are usually purchased rather than manufactured.
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