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    ELISA

    Uploaded by

    Abdul Kabir
    The document discusses several types of enzyme-linked immunosorbent assays (ELISA), including direct ELISA, indirect ELISA, and sandwich ELISA. Direct ELISA involves binding antigen to a substrate, adding a test sample containing antibody, then adding an enzyme-linked secondary antibody. Indirect ELISA involves binding antigen, adding sera with primary antibody, then adding conjugated antibody to detect primary antibodies. Sandwich ELISA measures antigen between two antibody layers (capture and detection antibody).

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    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd

    ELISA

    Uploaded by

    Abdul Kabir
    9 views2 pages
    The document discusses several types of enzyme-linked immunosorbent assays (ELISA), including direct ELISA, indirect ELISA, and sandwich ELISA. Direct ELISA involves binding antigen to a substrate, adding a test sample containing antibody, then adding an enzyme-linked secondary antibody. Indirect ELISA involves binding antigen, adding sera with primary antibody, then adding conjugated antibody to detect primary antibodies. Sandwich ELISA measures antigen between two antibody layers (capture and detection antibody).

    Original Description:

    ELISA ITS TYPES USAGES EXAMPLES DEFINITIONS

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    The document discusses several types of enzyme-linked immunosorbent assays (ELISA), including direct ELISA, indirect ELISA, and sandwich ELISA. Direct ELISA involves binding antigen to a substrate, adding a test sample containing antibody, then adding an enzyme-linked secondary antibody. Indirect ELISA involves binding antigen, adding sera with primary antibody, then adding conjugated antibody to detect primary antibodies. Sandwich ELISA measures antigen between two antibody layers (capture and detection antibody).

    Copyright:

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

    ELISA

    Uploaded by

    Abdul Kabir
    The document discusses several types of enzyme-linked immunosorbent assays (ELISA), including direct ELISA, indirect ELISA, and sandwich ELISA. Direct ELISA involves binding antigen to a substrate, adding a test sample containing antibody, then adding an enzyme-linked secondary antibody. Indirect ELISA involves binding antigen, adding sera with primary antibody, then adding conjugated antibody to detect primary antibodies. Sandwich ELISA measures antigen between two antibody layers (capture and detection antibody).

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    of 2

    Enzyme linked immunosorbent assay

    The enzyme-linked immunosorbent assay (ELISA) is an immunological assay commonly used to


    measure antibodies, antigens, proteins and glycoproteins in biological samples.

    Types

    1. Direct ELISA 2. Indirect ELISA 3. Sandwich ELISA 4. Competitive ELISA

    Direct ELISA

    Antigen is bound to the surface of the slide

    The test sample is poured on the substrate, if it contains the complimentary antibody then it
    binds with the antibody

    Following this rinsing is performed to remove unbound antibody

    Next, the mixture containing a secondary antibody with bound flourphore (Enzyme) is added,
    this antibody binds to the antibody antigen complex.

    Following this rinsing is performed to remove any unbound antibody.

    Finally substrate of the enzyme is added which changes color on reaction with enzyme thereby
    indicating a positive result.

    Indirect ELISA

    1. Antigen is bound on the polystyrene surface

    2. Sera samples are added to the wells

    3. Negative and positive controls are important parts of a test

    4. The wells are covered to allow antigen antibody in the incubation step.

    5. Wash buffer is added to remove any unbound antibodies.

    6. Now conjugated antibody (horse radish peroxide) is added which helps to detect and quantify
    the primary antibodies

    7. Rinsing is carried out using wash buffer

    8. Substrate solution (TMB) is added to each well and incubated


    Sandwich ELISA

    A sandwich ELISA measures antigen between two layers of antibodies (capture and detection
    antibody). The target antigen must contain at least two antigenic sites capable of binding to
    antibodies.

    Western Blot
    Widely used technique for detection and analysis of proteins.

    Blot is a membrane on which nucleic acids and proteins are immobilized.

    Process of transferring macromolecules from gel to membrane followed by its detection of


    these on the membrane is known as blotting.

    5 main steps

    Protein gel electrophoresis (Separation and characterization of proteins)

    Protein transfer

    Blocking

    Antibody probing

    Detection

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