An ELISA assay is only as good as the plate it runs on. The success of your experiment hinges on the quality of your plate and its ability to obtain accurate and reproducible results. Understanding how to select the best plate for your ELISA assay and their associated applications, you can best optimize your assay and ensure experimental success.
What Type of Plate Do Most Researchers Use?
Most researchers perform ELISA with microplates that have been constructed from polystyrene. This type of material is inexpensive and allows for the binding of a variety of molecules, including proteins and DNA. The material is also highly hydrophobic, meaning that it readily absorbs water and is relatively inert. This makes it an ideal surface for performing ELISA and immobilizing molecules.
How are Plates Coated?
Plate coating is the process that modifies the plates to allow for the binding of molecules of interest. This is typically achieved with a coating buffer, which is a solution that is pH-adjusted and contains high levels of salt. The buffer is applied to the plate, allowing for the desired molecule’s adsorption.
The process of plate coating occurs through the passive adsorption of the molecule onto the surface of the plate. This method is favored over covalent attachment, as it is less likely to damage the target molecule. Additionally, this approach allows for a high degree of flexibility; new molecules can be easily adsorbed onto the plate surface without the need for covalent modification.
What Variations in Plate Coatings Can Impact Binding?
The type of plate coating and the buffer used to coat the plate can impact the binding of molecules. For example, when using a coating buffer that is high in salt, the buffer will compete with the target molecule for binding sites on the plate surface. Additionally, certain coatings can sterically hinder the binding of molecules. By understanding the effects of plate coatings on binding, you can select a plate that will best suit your assay and desired outcomes.
What are the Benefits of Using Pre-Coated Plates?
Pre-coated plates offer several benefits that uncoated plates cannot. Using a pre-coated plate eliminates the need to coat your own plates, saving time and ensuring reproducibility. Additionally, pre-coated plates are often coated with high quality buffers that have been optimized for ELISA. This can result in improved assay performance and increased experimental success.
There are four main types of pre-coated plates: modified polymer surfaces, antibody-binding plates, biotin-binding plates, and fusion-tag binding plates. Each type of plate has its own unique benefits and applications.
- Modified Polymer Surfaces: Modified polymer surfaces are the most common type of pre-coated plate. They are coated with a variety of molecules that can improve experimental success, including antibodies, lectins, and proteins.
- Antibody-Binding Plates: Antibody-binding plates are coated with antibodies that selectively bind to a particular antigen. This type of plate is ideal for ELISAs that require a high degree of specificity.
- Biotin-Binding Plates: Biotin-binding plates are coated with biotin, an essential vitamin for human health. Biotin is a small, water-soluble molecule that binds to proteins that contain the biotin-binding site. This type of plate is ideal for ELISAs that require a high degree of sensitivity. These are also excellent for adsorption to polystyrene in situations where the target molecule is damaged by direct adsorption.
- Fusion-Tag Binding Plates: Fusion-tag binding plates are coated with fusion proteins that bind to a particular tag. The fusion-tag binding plate can be used to detect the presence of the target molecule in a sample. The plate is typically used in conjunction with a detection antibody that will bind to the target molecule and generate a signal that the reader can detect.
By understanding the different types of pre-coated plates available, you can select the plate that best suits your assay needs. With a wide variety of plates available, there is sure to be a plate that will meet your needs and help you achieve successful results.
Sources:
https://www.sciencedirect.com/topics/engineering/polymer-surface-modification
