This is the first of a series of posts on pros and cons of classical covalent links, which we will publish during the next few weeks. The very first one is about 3 well-known classical covalent links: carboxyl, hydroxyl and amino groups. In the next posts we are going to review other examples, such as tosyl or streptavidin beads.
Attaching a protein to a bead can be a detailed process that requires forethought and careful planning. Generally, a molecule is attached to a particle through a surface group available on the coating of that particle, for example in the case of streptavidin beads. In cases where the attachment is covalent, it is essential to choose a binding site on the molecule that will allow for proper orientation, maximally presenting the desired site to the sample while still retaining a strong attachment between the molecule and the bead.
Preparing magnetic beads for a particular assay, such as streptavidin beads, requires the beads to be functionalized. The beads need to be attached to the biological material that will serve as a capture molecule in the application. The particular type of attachment by which a molecule is linked to the bead will depend primarily on two things: the molecule being bound and the aim of the process.
Should I use streptavidin beads or gold particles when developing my IVD kits? In order to ensure the success of a protocol, it is essential to have a clear and unbiased knowledge base and a reliable source of reference material. When trying to decide the best platform or application to use for a process, it is critical to ensure that the information on which the decision will be based is generic and factual, and not propagated as promotional data.
There are a number of reasons why a lab might want to switch from colloidal gold particles to magnetic beads (for example, streptavidin beads) in an in vitro diagnostic assay, such as swapping from optical to magnetic detection in a lateral flow format or charging the format from lateral flor to a lab automated platform. Making the shift from utilizing one nanoparticle to another, however, is not a straightforward process. There are several areas where problems may arise. There is a considerable difference, for instance, in the way gold particles and magnetic beads bind a molecule. Moreover, the methodology for applications utilizing gold particles is significantly different from that of processes that make use of magnetic beads.
In vitro diagnostic (IVD) applications frequently make use of nanoparticles as solid-phase carriers for a given capture molecule. In order to be utilized in this manner, nanoparticles must first be coated, thereby attaching the capture molecules to the particles. There are notable differences in the way that different types of particles are coated, and this will be a significant factor in the decision to utilize one particle over another in a given assay, for example, streptavidin beads or latex particles.
Because of their significantly increased surface area and their ease of manipulation, in vitro diagnostic (IVD) assays commonly make use of nanoparticles, typically utilizing them as solid surface carriers for capture molecules. There are a number of particles that can be utilized in IVD assays. White latex particles, for instance, have traditionally been used in diagnostic tests such as immunoturbidimetry and nephelometry assays, but we find other interesting examples such as streptavidin beads.
Chemiluminescent Immunoassays (CLIAs) are excellent assays for high throughput, low analyte concentration, time sensitive testing and isolation. Using magnetic beads as the reagent in a CLIA, for example coated in the form of streptavidin beads, is an easy and established technique favored among many clinical scientists.
Magnetic bead suppliers vary in how they produce their product, how they deliver their product and how they guarantee their product. Some suppliers will also only provide common types of magnetic beads, such as magnetic streptavidin beads, while others will offer more specific products.
Non-specific background and auto-aggregation in chemiluminescent immunoassays are often caused by the presence of exposed hydrophobic surfaces on the magnetic beads. The use of a blocking reagents combined with gentle homogenous bio-magnetic separation will help reduce background and auto-aggregation of your coated beads.