What is Sandwich ELISA
The Sandwich Enzyme-Linked Immunosorbent Assay (ELISA) is a veritable powerhouse in the realm of medical research and diagnostics. This technique employs two antibodies, one of which is immobilized on a solid surface, while the other is labeled with an enzyme. The principle behind this assay is to first capture the target protein of interest using a solid phase antibody, and then detect the bound protein via a labeled detection antibody.
Typically, the solid phase antibody is coated onto the surface of a microtiter plate, while the detection antibody is labeled with an enzyme, such as horseradish peroxidase (HRP) or alkaline phosphatase (AP). In practice, the sandwich ELISA commences with the addition of the sample containing the protein of interest to the coated microtiter plate. This is then followed by washing to rid the plate of any unbound material, thus leaving only the protein of interest bound to the immobilized antibody.
Next, the labeled detection antibody is introduced into the plate and allowed to bind to the captured protein. After yet another washing step, a substrate for the enzyme is added, which elicits a color change in the presence of the enzyme. The intensity of the color produced is directly proportional to the amount of protein present in the sample.
The sandwich ELISA boasts a litany of advantages over other immunoassays. For one, it is highly specific, meaning it can detect only the protein of interest in a complex mixture of proteins with great accuracy. It is also highly sensitive, with a detection limit as low as a few picograms per milliliter of sample. Finally, it is relatively simple and inexpensive, rendering it a popular choice for a wide range of applications.
Figure 1. The sandwich ELISA.
The sandwich ELISA has several uses, including the diagnosis of illnesses. It can be used to detect particular diseases by identifying particular proteins in blood or other physiological fluids. For instance, the detection of troponin in blood can be used to diagnose a heart attack whereas the detection of prostate-specific antigen (PSA) in blood can be used to diagnose prostate cancer.
The sphere of biotechnology is where the sandwich ELISA has another significant use. This method can be used to estimate the amount of a certain protein that a cell or organism produces. This has enormous potential for improving protein synthesis procedures or researching the control of gene expression. It may also be used to discover novel protein-protein interactions, which is useful for deciphering the mechanisms behind diverse biological processes.
Despite its many advantages, the sandwich ELISA does have some limitations. One of the main limitations is that it requires two antibodies that recognize different epitopes on the protein of interest with high specificity. If suitable antibodies are not available, the technique cannot be used. Additionally, the binding of the protein of interest to the solid phase antibody may be impacted by the conditions used for coating the microtiter plate, which could affect the accuracy and reproducibility of the results.
In conclusion, the sandwich ELISA is a powerful and versatile technique for the detection and quantification of specific proteins in biological samples. Its high specificity, sensitivity, and relative simplicity make it a popular choice for a wide range of applications, including disease diagnosis and biotechnology research. However, its reliance on specific antibodies and potential issues with plate coating must be taken into account when designing and interpreting experiments.
