Antigens stimulate the body’s immune response by binding to antibodies and activating lymphocytes. Immunologists can utilize the unique binding properties between antigens and antibodies to identify specific cell types, track cellular behavior, and isolate or separate target cells within a biological sample.
Antigens are molecules that bind to specific antibodies or receptors on the surface of lymphocytes, and range in composition from proteins to lipids, polysaccharides, or other biomolecular substances. Antigen structure includes regions called “antigenic determinants,” or epitopes, that fit into the specific receptor or binding site.
Antigens play a vital role in triggering the body’s immune response. Antigens found on the surface of cells fit into antigen-binding sites on the paratopes of antibodies or into T- or B-cell receptors on the surface of lymphocytes.
Antigens and antibodies are distinct and complementary biomolecules. Antibodies are proteins that function in the immune system to identify or neutralize hostile pathogens by binding to their antigens. The tips of Y-shaped antibodies feature specialized antigen-binding sites called paratopes, into which the epitopes of antigens fit like a key into a lock.
“Pathogen” describes any bacterium, virus, parasite, fungus, or biocompound that can cause disease. Often, pathogens produce or present unique molecular structures that can act as an “antigen”. While antigens present on the surface of native cells go unnoticed by the immune system, antigens produced by pathogens trigger the body’s immune response.
There are three types of antigens, classified based on where they are produced. Self-antigens, or autoantigens, are produced in the body’s own cells; endogenous antigens are produced in intracellular bacteria or viruses; exogenous antigens are produced outside the body and are foreign to the immune system. The immune system relies on antibodies or receptor sites on T or B Cells to bind with foreign antigens to detect, track, and destroy pathogens present in the body.
B cells and T cells undergo a process called “central tolerance” or “negative selection,” in which cells with antigen-binding sites that might bind to or become activated by self-antigens are eliminated during development. Central tolerance ensures that the body can distinguish harmless self-antigens from the threats posed by foreign antigens, and healthy, functioning immune systems typically leave self-antigens alone.
As pathogenic cells invade the body, the foreign antigens they present bind with free antibodies, the antibodies in B Cell Receptors, or antigen-binding sites on T Cells. The antibody-antigen binding reaction may neutralize the pathogenic properties of the antigen or serve to “flag” the pathogen with antibodies that other elements of the immune system can target. Binding to a B Cell Receptor triggers the B Cell to differentiate into a plasma cell that produces antibodies that correspond to the foreign antigen, as well as secrete cytokines that influence how other immune cells function. In kind, binding to a T Cell Receptor induces the T Cell to differentiate into cells that can kill the pathogen or release cytokines to signal for broader immune response.
Antigens have come to play a useful role in cell sorting or cell separation procedures in laboratory research. Immunologists can mimic the mechanisms with which the body recognizes and targets foreign antigens, and use the selective properties of the antibody-antigen to identify cell types presenting specific antigens within a blood or tissue sample. These target cells can be separated from the rest of the sample, leaving cells that don’t present those antigens untouched.
The Buoyancy Activated Cell Sorting [BACS] procedures developed at Akadeum Life Sciences utilize the antibody-antigen reaction to perform accurate and efficient cell separation protocols. BACS methodology involves mixing antibodies with a cell suspension and using microbubbles to gently remove unwanted cells from the sample population.
To perform BACS with microbubbles, researchers utilize the kit configured to enrich the cell sub-type of interest. This kit contains all necessary reagents including streptavidin-coated microbubbles and a cocktail of biotinylated antibodies specific to the unwanted cell populations. The biotinylated antibodies with paratopes corresponding to the epitopes on the unwanted cells are introduced to the sample. These antibodies bind to the antigens, serving to identify the target cells to be removed. Next, streptavidin-coated microbubbles are mixed into the sample. The streptavidin on the microbubbles binds to the biotin on the antibodies that are attached to the antigens on the target cells. Finally, the buoyant microbubbles and their attached target cells gently rise to the surface of the sample to be removed, leaving the desired cells untouched and ready for analysis.
The selectivity of the antibody-antigen coupling, as well as the incomparable strength of the bonds between streptavidin and biotin in the streptavidin-biotin complex, make BACS with streptavidin microbubbles a compelling cell isolation method. In one hand, the biotinylated antibodies bind to only those cells in a sample that present the specific antigens with epitopes capable of binding to their paratopes. The unique binding correspondence between antibodies and antigens allows for precise labeling of only unwanted target cells, ignoring cell types that don’t feature these antigens. In another hand, the strong binding affinity between biotin and streptavidin ensures that these target cells remain securely adhered to the streptavidin-coated microbubbles as they float to the surface of the sample for collection and removal.
Traditional methods of cell sorting, isolation, or separation such as Fluorescence Activated Cell Sorting [FACS] or Magnetic Activated Cell Sorting [MACS] subject cells in a blood sample to intense mechanical or magnetic forces that can risk damaging healthy cells in the sample. Moreover, these methods rely on expensive equipment that can require specialized training to operate, and may be especially cost- and time-prohibitive for small labs or research facilities. That said, BACS protocols with biotinylated antibodies and streptavidin microbubbles are affordable, easy to use, and utilize only the gentle mechanics of buoyancy to isolate cells—posing little risk to the viability of cells in the sample. BACS is proven to deliver fast, scalable, and accurate results, yielding a pure sample of highly viable cells for analysis or downstream processing.
Learn more about Akadeum’s breakthrough BACS methodology and the efficacy of our wide offering of BACS products prepared for sorting a range of human and mouse cells. Use our precise antigen-antibody binding streptavidin microbubbles for high accuracy cell separation. Check out our app note and applications page to see how Akadeum’s BACS and microbubble technology is being used around the world.