What Is an Enzyme-Linked Immunosorbent Assay (ELISA)?
What Is an Enzyme-Linked Immunosorbent Assay (ELISA)?
Enzyme-linked immunosorbent assays (ELISA’s) are plate-based enzyme assays used for identifying and quantifying sample proteins, peptides, antigens, and antibodies that are targets in the research of a specific compound. The identification of a specifically targeted protein or antibody in enzyme-linked immunosorbent assays is completed by using antibodies to capture and immobilize the protein or antibody of interest and detect the presence of the targeted protein. The general ELISA applications are often referred to as a sandwich ELISA because the measured analyte is between two antibodies. One of the antibodies serves as the capture antibody, while the other is the detection antibody. Visit Surmodics IVD’s Sandwich ELISA Protocol
Why is ELISA Important?
Curing disease starts at diagnosis. It is imperative that diagnostic assays, including ELISA Assays be consistent and reliable to accurately detect disease. Developing accurate and reliable diagnostic assays starts with using the right reagents. Surmodics IVD’s reagents include the industry’s gold standard in protein stabilizers/blocking buffers designed to increase sensitivity and specificity while decreasing non-specific binding. We also offer best-in-class detection substrates for color (Colorimetric Substrates such as TMB, ABTS, PNPS, BCIB/NBT) and light (Chemiluminescent Substrate such as Luminol) development as well as a selection of accessory reagents including our high-performance diluents designed to reduce matrix interferences including Human Anti-Mouse Antibodies (HAMA) and Rheumatoid Factor (RF) interferences.
We also manufacture and supply a line of microarray slides and surface modification technologies as well as supply high quality DIARECT™ antigens and antibodies from our partner company, BBI™ Solutions.
Basic ELISA Principals
An enzyme-linked immunosorbent assay (ELISA) is developed by using a specific capture antibody which is used to coat a microtiter plate. After the sample is added, a primary antibody will bind the target antigen or protein of interest. A secondary conjugated detection antibody will then bind to the primary antibody. Reagents that are unbound are removed through washing, along with the unbound antibody. Once a substrate is added, a colorimetric reaction develops proportional to the protein concentration that is bound. The concentration of the target protein in the sample is determined by comparing to a known concentration of the protein in the standard curve.
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An ELISA Test – How to Run an ELISA Test
Enzyme-linked immunosorbent assay (ELISA) is a method used to identify and quantify substances such as proteins, peptides, antibodies, hormones, and other analytes. ELISA kits have become one of the most used assay techniques for research laboratories, clinical diagnostics companies and more. ELISA is used in other formats such as testing individual samples and automated screening. ELISA is used as a common diagnostic tool for medicine, biotechnology, plant pathology, and quality assurance in many industries.
An assay is completed using a specific antibody which is used to coat a microtiter plate. Detection antibodies are used for reporting the quantification of the ELISA. After the sample is added, a primary antibody will bind the target protein of interest. A secondary conjugated detection antibody will then bind to the primary antibody Reagents that are unbound are removed through washing. Once a substrate is added, a color reaction develops proportional to the protein concentration that is bound. The concentration of the target protein in the sample is determined by comparing to a known concentration of the protein in the standard curve.
The high sensitivity ELISA technology depends on the affinity of the antibodies as well as the system one is using. A standard range for the assay shows the upper and lower limits of analyte concentration. The accuracy of the ELISA is calibrated with an external reference. Standards can be found available from the National Institute of Biological Standards and Control.
Visit our ELISA Assay Solutions and Protocol to learn more:
Different Types of ELISA
ELISA Assays and Applications
The main enzyme-linked immunosorbent assay subtypes include direct and indirect sandwich ELISA, and competitive ELISA also known as an inhibition ELISA. The type of enzyme-linked immunosorbent assay used varies by the objective of the developer and how the antibodies and analytes are bonded. Surmodics IVD proudly offers several immunoassay components for both competitive ELISA formats and sandwich ELISA formats.
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Frequently Asked Questions:
What Can ELISA Assay be Used for?:
Enzyme-linked immunosorbent assays have become a widely used assay technique for research laboratories, clinical diagnostics companies and more. ELISA’s are used in formats such as assaying individual samples, automated screening, and is used as a common diagnostic tool for medicine, biotechnology, plant pathology, and quality assurance in many fields.
ELISA methods measure analytes in specific biological samples to diagnose current or past infections, diseases, allergies, and more. ELISA Assays that utilize biological samples include HIV infection Assays, Pregnancy Assays, Lyme Disease Assays, COVID-19 Assays and more, using the measurement of soluble indicators in a saliva sample, serum or blood sample, or other bodily fluid or tissue sample.
Accurate diagnosis is imperative for providing proper care and treatment. Because Surmodics IVD’s reagents are manufactured in an ISO 13485:2016 & 9001:2015 facility, ELISA developers can rest assured that lot-to-lot consistency, reproducibility, and optimal performance is achieved.
Visit the link below to learn more about Surmodics IVD’s quality standards:
What to Expect During an ELISA Assay:
For the patient, the process of an enzyme-linked immunosorbent assay is typically simple and quick. Medical personnel will have blood drawn or other bodily fluid or tissue obtained from the patient. An ELISA kit will be used to assay the patient’s sample and indicate a reaction, or lack thereof. The laboratory assay will either be positive, negative, or invalid.
If cross-reactivity occurs during the development of an assay this can impact the accuracy of the assay results, leading to false positives. Developers can overcome false positives by utilizing Surmodics IVD’s Protein Stabilizers, Blockers and Diluents.
What is the most common ELISA Test?
Sandwich ELISA assays are the most commonly used assay test. In a sandwich ELISA assay, two specific antibodies sandwich an antigen or antibody of interest, known as matched antibody pairs. The capture antibody is coated on a microplate. Target proteins from the sample bind to the captured antibody if present. A conjugated antibody binds to detect the target protein when added. Then an ELISA substrate is added, producing a signal proportional to the given analyte in a sample. Although these ELISA tests are highly specific assays, they often need additional optimization due to a risk for false positive results.
Surmodics IVD’s Assay Diluents, MatrixGuard™ Diluent and Surmodics Assay Diluent (Protein-Free) are the gold standard for reducing false positives in your assay. The two formulations provide different blocking strategies to reduce or eliminate matrix interferences, while maintaining the clinical utility of the assay.
To learn more and/or request free samples for evaluation, contact our team today!
