Researchers have traditionally used agarose beads for immunoprecipitation (IP). However, there has been a growing trend in recent years favoring the use of magnetic beads. According to a recent survey of 1,013 scientists, 60% of the respondents will start using magnetic technology in the next three years.
So, what makes magnetic beads an attractive alternative to traditional agarose beads? Read the rest of this article to learn why most laboratories now choose magnetic beads for IP.
Magnetic Beads and Their Key Features: Taking a Closer Look
The use of Protein A, Protein G or Protein A/G magnetic beads in IP has been gaining popularity due to a number of reasons. For one, studies show that magnetic beads exhibit a faster rate of protein binding, and offer reduced antibody consumption and sample loss. Magnetic beads also exhibit low nonspecific binding, and optimized IgG binding capacity. Additionally, many laboratories switched to magnetic beads since they produce cleaner, more consistent results in significantly less time.
High Binding Capacity. While agarose beads may have a porous center which significantly increases their binding capacity, magnetic beads are significantly smaller than agarose beads (1 to 4μm). This gives them an effective surface area-to-volume ratio for optimum antibody binding. In addition, magnetic beads can aggregate without the need for centrifugation, thereby increasing the yield of delicately attached protein complexes.
Reduced Antibody Consumption. Since agarose beads are porous and have high binding capacity, they require larger amounts of antibodies to produce accurate results. The antibody can be trapped inside the bead and fail to properly bind the protein of interest. When this happens, you may need to use more antibody. You wouldn't have this problem with magnetic beads since they are non-porous and antibody binding is limited to the outer surface of the bead.
Keep in mind that when the amount of antibody available for the immunoprecipitation experiment is less than sufficient to saturate the agarose beads, you can end up with particles that are only partially coated with antibodies. This can be a problem since the unsaturated portion of the beads will then be free to bind with anything that will stick. In such cases, you can expect elevated background signal due to non-specific binding of lysate components to the beads.
Reduced Sample Loss. Since magnetic beads do not require centrifugation, there is no risk of aspirating immune complexes that are bounded to the beads. This also reduces the risk of breaking weak antibody-antigen binding and the subsequent loss of target protein for a more accurate quantitation of your protein of interest and better reproducibility.
Fast, Easy and Convenient Separation Procedures. Agarose beads and magnetic beads both bind to the Fc region of IgG in the process of isolating specific proteins and protein complexes from cell lysate. However, since magnetic beads do not have to be collected through centrifugation (they are collected using a magnetic device close to your tube), the separation process can be completed in 30 minutes. This allows for the removal of unwanted supernatant without disturbing your protein pellet, improves the capture of transient protein complexes and reduces background noise.
Price. Admittedly, magnetic beads are more expensive than agarose beads. However, when you consider the fact that there is no minimum quantity of magnetic beads required and that you can use considerably less of these beads in your experiment, then you can see that magnetic beads may be more competitively priced as compared to agarose beads.
Equipment. While the high-power magnets that are required for magnetic bead-based IP reactions may be cost prohibitive, using magnetic beads allow you to complete your experiments and generate more data in a shorter amount of time. Additionally, automated immunoprecipitation devices are now becoming more readily available so you can significantly reduce the amount of work and time required to perform an IP, and use them for high-throughput applications as well.
