Radiolabeling of biomolecules


Radiolabeled biomolecules, including peptides and antibodies with short biologic half-lives, are gaining increasing importance for PET imaging studies of receptor expression and activity in living subjects. Because of the fragility of biological molecules (especially proteins), then cannot survive high temperatures, organic solvents, or strong acid or basic conditions that are often used to incorporate radioisotopes into small molecules. The labeling must be done under mild conditions (e.g. ambient temperature up to 40C, aqueous media, etc.)

Biological molecules sometimes labeled with linkers that can quickly reactor under mild conditions with the radioisotope. For example, numerous chelators have been designed to quickly and stably chelate a variety of positron-emitting radiometals. The biomolecule can be labeled with the linker first, in which case the reaction and purification process need only be mild -- not necessarily fast. The second step (chelation), however, must be fast. Alternatively, the radiometal can first be chelated (e.g. if high temperatures are needed for rapid chelation), and then the chelating group (with radiometal) rapidly coupled to the biomolecule.

Due to the high availability and excellent properties of fluorine-18, there is extensive interest in labeling biomolecules with this isotope. Because fluorine-18 is almost always incorporated into molecules under harsh conditions, a prosthetic group strategy is employed. First the fluorine-18 is incorporated under harsh conditions into a small molecule (the "prosthetic group"), and this prosthetic group is then rapidly conjugated to the biomolecule under rapid and mild conditions. The biomolecule may need to be modified with a linker to become reactive with the prosthetic group.

By clever design of the biomolecule, it is possible to perform site-specific conjugation, to ensure that conjugation does not occur at a site that could interfere with the biological function of the molecule. One way to do this is to use a prosthetic group that binds to a functional group that is rare or unique in the engineered biomolecule. For example, thiol is an uncommon functional group in peptides and proteins, and by incorporating cysteine at the location where labeled is desired. This site can be reacted with, e.g., a maleimide-containing prosthetic group. Recent research in this field has focused on developing a variety of fast and specific labeling strategies.

An example of a non-specific prosthetic group is N-succinimidyl-4-[F-18]fluorobenzoate ([F-18]SFB).

Suggested Reading

The following review paper covers various F-18-labeling methods and compares commonly used prosthetic groups:

  • Richter, S., Wuest, F., 2014. 18F-Labeled Peptides: The Future Is Bright. Mol. Basel Switz. 19, 20536–20556. doi:10.3390/molecules191220536 (Journal Link)