ELISA Kits Preparation & Handling Tips

Introduction to ELISA kits and their importance

The Enzyme-linked immunosorbent assay (ELISA) is one of the most commonly used techniques to measure the presence of a soluble analyte, like cytokines or chemokines, in serum, plasma, cell supernatant, and other biological samples. ELISA is an antigen-antibody interaction in which specific antibodies bind to the target antigen and detect the presence and quantity of antigens binding. Here, we are introducing two types of ELISA: traditional ELISA and AlphaLISA® on the basis of their workflow.

Traditional ELISAs are standard immunoassays that are typically performed in multiple-well microplates, coated with capture antibodies on the microtiter well, to immobilize soluble proteins. Liquid sample containing the target analyte is added to a microplate well that is coated with capture antibodies. Any target analyte in the sample binds to the antibodies. The bound proteins are then recognized by a second antibody, known as the detection antibody, which is often labelled with an enzyme. In some cases, the detection antibody is biotinylated, and the samples are incubated with a streptavidin-enzyme conjugate.

Finally, a substrate is added which changes its color when it interacts with the enzymes linked to the analytes or antibodies. The color change is proportional to the amount of antigen captured. By using a plate reader/spectrophotometer to measure this signal and plotting the resulting values against a standard curve with known quantities of analyte, you can obtain precise, quantitative measurements.

A typical sandwich ELISA workflow can take at least 22 hours as it includes coating the microplate with antibodies overnight, many washing steps, time-sensitive incubation, and plate shaking before the assay is quantitated by running against a standard curve. Genomax distributes two types of ELISA kits to streamline this workflow, thereby reducing the number of protocol steps and saving you time. 

AlphaLISA is a bead-based, no-wash assay technology that is used to study biomolecular interactions in a microplate format in less than 3 hours for most applications and uses small sample volumes (1-5 µL) to conserve precious sample. AlphaLISA assay is easy to run and it is a homogeneous assay without wash steps. In an AlphaLISA assay, Alpha Donor and Acceptor beads are brought together through binding to the target. The donor beads are excited with red light from a microplate reader, and a cascade of chemical reactions is set in motion, creating an amplified signal. A singlet oxygen is generated by the donor beads, which excites the acceptor beads and will emit light proportional to the level of interaction. The assay can be quantitated by running against a standard curve.

Pre-assay preparation of ELISA kits

For increased flexibility and convenience, we offer four types of ELISA kits (from BioLegend), including RAPID MAX™ kits that cut your assay time down to less than 90 minutes, easy-to-use LEGEND MAX™ kits containing pre-coated plates, and economical sets with the required components to develop your own ELISA. Each of our ELISA kits are designed for the quantification of proteins in biological samples including cell culture supernatant, serum, and plasma. Our kits offer high accuracy, sample linearity, and testing on biologically relevant samples.

For maximum flexibility in studying biomolecular interactions, from studying pathways or phosphorylated proteins in whole cells, or studying endogenous receptors, measuring target engagement in cells, studying fusion-tagged proteins or protein-protein interactions (PPIs), or when running large numbers of ELISA assays from the same sample, we offer the AlphaLISA Assays to expand your research possibilities.

Handling and storage of ELISA kits
Each of the ELISA kits is fully validated and contains a Technical Data Sheet to explain the product information, kit contents/ materials provided, and storage information of kit components.

Shelf life and expiration of ELISA kits

When you are ready to use the ELISA kit, take the reagents out and prepare them by following the protocol/ Technical Datasheet provided with the kit. Unused thawed reagents can be stored and conserved for future use. Refer to the Technical Datasheet for recommended storage conditions and corresponding shelf life. Do not use the ELISA kits beyond its expiration date.

Tips for maintaining the quality and accuracy of ELISA kits

Please read the Technical Datasheet provided with the kit before use. Bring all reagents to room temperature prior to use. After reconstitution of the lyophilized standard with Assay Diluent, aliquot into polypropylene tubes and store at recommended storage conditions as per the Technical Datasheet for future experiments. Avoid freeze-thaw cycles of the reconstituted standard as loss of activity may occur.

Conclusion and summary of key points

Genomax distributes BioLegend’s and Revvity’s ELISA kits and sets, microplates, buffers, and controls for two types of ELISA: traditional ELISA and AlphaLISA®. Find all the reagents for your ELISA workflow with our economical kits, antibodies, ancillary reagents, and standards from BioLegend. For flexibility to measure even large biomolecules (up to 2000 kDa) and very weak interactions (mM), and the ease of running the assay with a simple workflow with no wash steps, choose the AlphaLISA assay kits for your application.

Looking for additional resources or need help with troubleshooting your experiment? Contact us at info@genomax.com.sg with additional questions.

Leave a Reply

Your email address will not be published. Required fields are marked *

Contact Us
close slider