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    Hepatitis B Surface Antigen

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    selam kalayu
    This document describes the sandwich ELISA method for detecting hepatitis B surface antigen (HBsAg) in samples. It works by coating monoclonal antibodies on wells to capture HBsAg from samples. Another monoclonal antibody labeled with an enzyme is then added to form antibody-antigen-antibody sandwiches bound to the wells. Adding a substrate causes a color change if HBsAg is present, indicating a positive result. The procedure involves incubating samples and controls in wells, washing, adding substrate, and reading results colorimetrically to determine if samples are positive or negative for HBsAg infection.

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    © All Rights Reserved
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    Hepatitis B Surface Antigen

    Uploaded by

    selam kalayu
    402 views2 pages
    This document describes the sandwich ELISA method for detecting hepatitis B surface antigen (HBsAg) in samples. It works by coating monoclonal antibodies on wells to capture HBsAg from samples. Another monoclonal antibody labeled with an enzyme is then added to form antibody-antigen-antibody sandwiches bound to the wells. Adding a substrate causes a color change if HBsAg is present, indicating a positive result. The procedure involves incubating samples and controls in wells, washing, adding substrate, and reading results colorimetrically to determine if samples are positive or negative for HBsAg infection.

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    Hepatitis B surface antigen

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    This document describes the sandwich ELISA method for detecting hepatitis B surface antigen (HBsAg) in samples. It works by coating monoclonal antibodies on wells to capture HBsAg from samples. Another monoclonal antibody labeled with an enzyme is then added to form antibody-antigen-antibody sandwiches bound to the wells. Adding a substrate causes a color change if HBsAg is present, indicating a positive result. The procedure involves incubating samples and controls in wells, washing, adding substrate, and reading results colorimetrically to determine if samples are positive or negative for HBsAg infection.

    Copyright:

    © All Rights Reserved
    Download as DOCX, PDF, TXT or read online from Scribd
    Download as docx, pdf, or txt
    402 views2 pages

    Hepatitis B Surface Antigen

    Uploaded by

    selam kalayu
    This document describes the sandwich ELISA method for detecting hepatitis B surface antigen (HBsAg) in samples. It works by coating monoclonal antibodies on wells to capture HBsAg from samples. Another monoclonal antibody labeled with an enzyme is then added to form antibody-antigen-antibody sandwiches bound to the wells. Adding a substrate causes a color change if HBsAg is present, indicating a positive result. The procedure involves incubating samples and controls in wells, washing, adding substrate, and reading results colorimetrically to determine if samples are positive or negative for HBsAg infection.

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    Hepatitis B surface antigen (HBsAg) using

    Sandwich ELISA

    Principle
    HBsAg adopts the "sandwich principle" as the basis of the assay. When a positive
    sample is incubated into the well, the coated monclonal antibody (solid phase antibody)
    binds the HBsAg of the sample. A further incubation with a second monoclonal anti-
    HBsAg labelled with the enzyme peroxidase (anti-HBsAg-HRP) leads to the sandwich
    immunocomplex bound onto the well: solid phase antibody: HBsAg Antibody-HRP. The
    activity of the HRP is then revealed by the addition of the substrate, which becomes blue
    and then turns to yellow after blocking the reaction with acid.

    MATERIALS
     Samples 1, 2, 3
     HBsAg ELISA kit
     96-Well Microtiter Plates
     HRP conjugates
     Negative control
     Positive control
     Substrate A and B (TMB and H O )
    2 2
     Stop solution (H SO )
    2 4
     Washing solution
     Incubator

    Procedures
    1. Write down the relative numbers of specimens and wells on the
    data sheet. One well for the blank, additional wells for the
    controls and one well for each specimen.
    2. . Reserve one well for blank, Add one drop of samples, positive
    control and negative controls to each well.
    3. Add one drop of Anti-HBsAg peroxidase-conjugate working
    solution into each well.
    4. Cover the plate with adhesive plastic and incubate the plate in a
    +37°C incubator for 30 minutes.
    5. At the end of the incubation wash the strips 5 times with the
    working wash solution either manually or with an automatic
    washer. After the final washing, make sure that whole solution is
    perfectly removed from each well.
    6. Add one drop of Substrate solution A and B, including 1A. Mix
    horizontally and incubate for15 minutes at +37°C
    7. Stop the reaction by adding one drop of Blocking Reagent to
    each well.
    8. After adding the stop solution, read the color developed on the
    microplate reader.
    Results and analysis
    The amount of color intensity can be measured and it is proportional to the amount of
    antigen captured in the wells, and to its amount in the sample respectively. Wells
    containing samples negative for HBsAg remain colorless. Intensive yellow color
    develops in Positive control and HBsAg positive sample wells. If it’s positive that means
    you are infected by HBV and if its negative you are not.

    Name: Kassa Mengzemwork Kalayu

    Student id: 185019011

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