EIA Assay Procedure

Posted by Jack on November 23, 2022
Table of Contents


    The ELISA assay procedure is a process of detecting analytes in a sample by using an antibody that is specific to the antigen. The EIA can test for the presence of antibodies or antigens, depending on the type of test being conducted. The EIA technique has been used extensively for many years and is considered one of the most sensitive techniques available for detection of analytes.

    Study Population

    It is important to define the study population. The population is the group of people who are being studied. For example, if you’re conducting an EIA assay on children, then your sample will be comprised of children only. You should also specify their ages (e.g., 3-year-olds).

    Age range: It is important to specify whether you are looking at adults or children and what age range they fall under (e.g., 3-year-olds).

    Gender: Specify whether the study population consists of male or female participants only, or if it includes both genders equally.

    Ethnicity: If your studies include people from different ethnic backgrounds, it is essential that you specify which ones these are so that they can be included appropriately in future experiments with similar demographics

    Ethics Statement

    The IRB must approve the consent form used for the study.

    The IRB must approve the informed consent process.

    ELISA Procedure

    ELISA is a type of immunoassay used to detect and measure the presence of an analyte in a sample. ELISA stands for enzyme-linked immunosorbent assay, which means that it uses antibodies to bind to the analyte, then detects these bound antibodies with a secondary antibody conjugated to an enzyme. This second antibody can be tagged with a dye or other indicator molecule, along with enzymes that catalyze reactions resulting in color change or fluorescence.

    The appropriate assay technique must be used to detect analytes in the sample type.

    If you want to know more about assay techniques, check out this resource!

    It's important to select the appropriate assay technique for detecting your analyte. For example, if you want to detect a protein in a tissue sample, then immunohistochemistry (IHC) would be an appropriate choice. However, if you're looking for DNA originating from a virus or bacteria.in feces samples using polymerase chain reaction (PCR), then nested PCR would be more appropriate.


    Materials needed:

    • Specimen. Your blood or urine sample should be fresh and stored at room temperature, but not frozen. The specimen should be a minimum of 5 ml and a maximum of 10 ml in volume. If you're testing your blood, it helps to first use a finger prick to draw some of your own blood into an empty sample tube (we recommend using DTF Plus Red Blood Cell collection tubes). Then add one drop of anticoagulant (e.g., sodium citrate) and mix well by gently rolling the tube between your hands for several seconds prior to adding an additional 10 ml anticoagulant to bring the total volume up to 10 ml (this will prevent clotting).

    Antibody. This works as an indicator that there is sufficient IgE present in your sample's antibodies for detection purposes; it also contains Biotin-XL™ reagent for labeling purpose later on when we're ready for testing


    The EIA assay procedure is as follows:

    • Prepare the sample solution by dissolving 1 ml of sample in 9 ml of PBS using a vortex mixer.
    • Prepare reagent solutions A and B at room temperature by adding 100 ml of reagent stock and 10 ml of diluent directly into 100-ml volumetric flasks, respectively. Note that each flask should contain approximately 1 g/L of protein or nitrophenol (i.e., 1010 moles/liter). Alternatively, these solutions can be prepared from concentrated stocks and diluted with diluents prior to use for improved stability over time if needed; however, this will require additional volumes during the assay itself because less accurate results will result from preparing them beforehand due to losses during handling steps such as pipetting small amounts when transferring volume changes due to evaporation rates during long periods between preparations

    Data Analysis

    • The data is analyzed to determine the concentration of the analyte in your sample. In other words, how much of that substance is present?
    • The analysis involves a curve fitting function, like one- or two-parameter approximations for nonlinear regression analysis and linear regression analysis, respectively. You might also see them referred to as "least squares" or "maximum likelihood".
    • This process involves finding an equation that best fits the experimental data points: it attempts to minimize errors between observed values and calculated values at each point on your graph.


    • Check the kit components.
    • Check the reagents.
    • Check the equipment.
    • Check the assay procedure.
    • Check the sample.
    • Check for any contamination or phenotypic change in your culture or cell lines, which may introduce errors into your results and can be difficult to detect with assays that measure protein levels by ELISA or Western blotting (protein quantification) methods.

    The EIA Assay is an immunoassay that uses an antibody-antigen reaction to determine the concentration of an analyte.

    The EIA Assay is an immunoassay that uses an antibody-antigen reaction to determine the concentration of an analyte. Antibodies are proteins produced by the immune system and they interact with antigens on foreign substances, such as bacteria. The antigen-antibody reaction occurs when the antibody binds to specific epitopes on the antigen's surface. This binding causes a change in color or fluorescence, which can be detected using spectrofluorometry or light scattering techniques. There are many advantages of using this method; however, there are also some disadvantages associated with it as well:


    • It can be used with any type of sample that contains antibodies against your target analyte
    • Less expensive than other methods like gas chromatography (GC) or mass spectrometry, making it ideal for many applications where cost is an important factor


    • Results may not always be accurate since they depend heavily on how accurately you calibrate your reagents before performing tests


    In conclusion, the ELISA assay is a useful technique for detecting analytes in blood, serum, plasma and other biological fluids. The method has been used as an alternative to radioimmunoassay (RIA) or fluorescence activated cell sorting (FACS). It is also an excellent tool for research studies involving small numbers of samples such as in clinical trials where there are fewer than 100 subjects involved.

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