Sophisticated affinity chromatography techniques can mitigate the frustrations and challenges of isolating important proteins that are all too often drowned out by the enormous amounts of others. Such is the dilemma of proteomics—capturing proteins involved in genetic regulation, cell signaling, disease development, and other noteworthy functions. Expressed at the lowest of levels, these proteins would remain hidden without the help of today’s affinity chromatography tools.
Isolating and purifying proteins has long been the ambition of researchers. As early as the 1930s, labs were using separation techniques resembling affinity chromatography. Over the years, scientists took advantage of the efficient binding of enzymes with specific inhibitors and used it as a springboard to improved methods. Progress in developing the technology reached such a height that in 1948 Sweden’s Arne Wilhelm Tiselius received the Nobel Prize in Chemistry for his work in perfecting affinity chromatography and developing numerous types of gels used for adsorption.
Whereas the pioneers had only a few protocols at their disposal, the market now offers a wide selection of products to assist you in studying all types of proteins. In addition to the crude polymers and resins of yesteryear, you can find specialized adsorbents, the solid substrates to which ligands are attached. Made of materials such as agarose, sephadex, and various derivatives of cellulose, just to name a few, the substrates now take the form of multiwell microplates, magnetic particles, spin columns—in addition to the original beads, which are still popular.
The type of substrate you choose will depend in part on the ligand, which is responsible for binding to the target protein. While polystyrene is especially well suited to protein ligands, magnetic particles are better at adsorbing certain antibodies and strepavidin, making magnetics an excellent choice for retrieving antigens, lectins, enzymes, and nucleic acids. In addition to traditional columns, miniature spin columns packed with gel are now available to isolate small amounts of proteins with the help of centrifugation.
Or, you may have to choose your substrate based upon the type of tag you’ve engineered into your target protein. These tags include calmodulin-binding peptide, strepavidin-friendly biotin, the cellulose-binding tag, the maltose-binding protein, and a short peptide of six histidines. For the popular histidine tags, you’ll be using a substrate of polymer-coated beads made of nickel, zinc, or cobalt, which have an affinity for histidine-tagged proteins.
With all kinds of substrates, tags, buffers, and reagents, the options are practically endless with today’s market. The range of affinity chromatography products, such as those below, give you the power to isolate proteins of nearly all sizes, shapes, and abundance.
Now, for the first time, you can identify and characterize high-value, low-abundant proteins faster and more efficiently without interferences. The Agilent Multiple Affinity Removal System allows you to remove unwanted, multiple high-abundant proteins from human serum, plasma, CSF samples and more — with just one device.
