Competitive ELISA Technique
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Questions and Answers

What is the primary purpose of immunoassay techniques?

  • To identify and quantify specific antigens in a sample (correct)
  • To destroy foreign biological materials
  • To manufacture immunoglobulins
  • To stimulate the production of antibodies in the body

    • What is the role of B-lymphocytes in the immune system?

  • To differentiate into T-lymphocytes
  • To produce immunoglobulins in the bone marrow
  • To destroy antigens directly
  • To produce antibodies in response to antigens (correct)

    • What is the term for the biochemical compound in a sample that is detected or analyzed in immunoassays?

  • Antigen
  • Immunoassay
  • Analyte (correct)
  • Antibody

    • What is the percentage of human glycoprotein antibodies that are in the immunoglobulin class IgG?

    <p>80%</p> Signup and view all the answers

    What is the shape of IgG antibodies?

    <p>Y-shaped</p> Signup and view all the answers

    What is the term for the part of the antigen that an antibody binds to?

    <p>Epitope</p> Signup and view all the answers

    What is the function of T-lymphocytes in the immune system?

    <p>To mark antigens-antibody complex to be destroyed</p> Signup and view all the answers

    What is the term for the large glycoprotein molecules produced by B-lymphocytes?

    <p>Antibodies</p> Signup and view all the answers

    What is the molecular weight of IgG molecules?

    <p>150,000 Daltons</p> Signup and view all the answers

    Where are the genes for the heavy chains of IgG molecules located?

    <p>Chromosome 14</p> Signup and view all the answers

    What type of bonds connect the heavy and light chains of IgG molecules?

    <p>Di-sulphide bonds</p> Signup and view all the answers

    What is the antigen binding site of an IgG molecule?

    <p>Determinant specific region</p> Signup and view all the answers

    What is an example of a naturally occurring antigen in the body?

    <p>HCG hormone</p> Signup and view all the answers

    What is the method used to produce large quantities of antibodies in the lab?

    <p>Injecting antigens into goats, sheep, or horses</p> Signup and view all the answers

    What is the characteristic of monoclonal antibodies?

    <p>They attach only to a specific antigen binding site</p> Signup and view all the answers

    What is the purpose of ELISA technique?

    <p>To detect the presence of an antibody or an antigen in a sample</p> Signup and view all the answers

    What is the purpose of an enzyme-conjugated anti-immunoglobulin in an ELISA?

    <p>To detect the antibodies being tested for</p> Signup and view all the answers

    What is the result of the reaction between the substrate and the enzyme in an ELISA?

    <p>A color change that is proportional to the amount of antigen bound</p> Signup and view all the answers

    What is the main difference between direct and indirect ELISA?

    <p>The sequence of antibody addition</p> Signup and view all the answers

    What is the purpose of the spectrophotometer in an ELISA?

    <p>To measure the intensity of the color produced</p> Signup and view all the answers

    What type of ELISA is used to detect very small analyte concentrations in a biological sample?

    <p>Sandwich ELISA</p> Signup and view all the answers

    What is the shape formed by the antibodies and the antigen in a sandwich ELISA?

    <p>Sandwich</p> Signup and view all the answers

    What is the advantage of using two antibodies in a sandwich ELISA?

    <p>It is highly specific, since two antibodies are required to bind to the protein of interest</p> Signup and view all the answers

    What is the purpose of the ELISA reader?

    <p>To measure the intensity of the color produced</p> Signup and view all the answers

    What is the main characteristic of a competitive ELISA?

    <p>Using a conjugated antigen to compete with the antigen present in the sample</p> Signup and view all the answers

    What happens when there is more antigen present in the sample in a competitive ELISA?

    <p>Less conjugated antigen will bind to the capture antibody</p> Signup and view all the answers

    What is the purpose of the first wells in a 96-well plate in a competitive ELISA?

    <p>To draw the standard curve</p> Signup and view all the answers

    Why is it important to read the kit instructions carefully before starting a competitive ELISA?

    <p>To avoid disturbance and loss of expensive kits</p> Signup and view all the answers

    What is the purpose of the quality control samples provided with the kit?

    <p>To test the performance of the assay</p> Signup and view all the answers

    How is the standard curve drawn in a competitive ELISA?

    <p>By plotting the concentration on the X-axis and the absorbance on the Y-axis</p> Signup and view all the answers

    What is the purpose of the standard curve in a competitive ELISA?

    <p>To determine the unknown concentration of each sample</p> Signup and view all the answers

    What is the characteristic of a positive result in a competitive ELISA?

    <p>The solution changes color</p> Signup and view all the answers

    Study Notes

    Immunoassay

    • Immunoassay is a technique that uses the binding between an antigen and its homologous antibody to identify and quantify specific antigens or antibodies in a sample.
    • Samples can be urine, saliva, tears, or any biochemical material.

    Antigens and Antibodies

    • Antigens are substances that stimulate the production of antibodies when they enter the body.
    • Antibodies are large glycoprotein molecules produced by B-lymphocytes in response to antigens.
    • Antibodies are designed to bind to specific surface binding sites or epitopes on the antigen.

    Structure of Antibodies

    • Over 80% of human glycoprotein antibodies are in the immunoglobulin class IgG.
    • IgG molecules have a molecular weight of 150,000 Daltons and are made of 2 long (heavy) chains and 2 short (light) chains connected by di-sulphide bonds.

    Antibody Production in Lab

    • Small quantities of antibody can be produced by injecting an antigen into small mammals, such as mice, rats, or rabbits.
    • Large quantities of antibody can be produced by injecting antigens into goats, sheep, or horses to obtain anti-sera with polyclonal antibodies.
    • Monoclonal antibodies can be produced by combining antibody-secreting B-lymphocytes with cancer cells (hybridomas).

    ELISA (Enzyme-Linked Immunosorbent Assay)

    • ELISA is a biochemical immunology technique used to detect the presence of an antibody or an antigen in a sample.
    • ELISA can be used to detect hormones, drugs, toxins, and genetic modified crops.

    Non-Competitive ELISA Requirements

    • Antigens (Ag) fixed to a solid surface (immobilized) on a 96-well plastic plate.
    • Antibodies (Ab) in solution to be tested.
    • Enzyme-conjugated Anti-immunoglobulin (Ab) against the antibodies being tested for.
    • Substrate binds to enzyme and produces color.

    ELISA Types

    • Direct ELISA: antigen is immobilized on a plate, and primary detection antibody is added.
    • Indirect ELISA: antigen is immobilized on a plate, primary detection antibody is added, and then a secondary antibody conjugate with the enzyme is added.
    • Sandwich ELISA: two specific antibodies are used, forming a sandwich shape with the antigen.
    • Competitive ELISA: a capture antibody is coated on a microplate, and a conjugated antigen is used to compete for binding with the antigen present in the sample.

    ELISA Equipment

    • ELISA reader
    • ELISA washer
    • Washer solutions
    • ELISA monitor
    • ELISA kits

    Analysis of Results

    • Classic method: read the results using a spectrophotometer, and draw the standard curve.
    • Soft ware method: specify the standard concentrations on the x-axis and the reading of each standard on the y-axis, and draw the standard curve.

    Results

    • The quality control sample concentration is determined from the standard curve.
    • Positive results are determined by the color change in the solution, and negative results are determined by the absence of color change.

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    Related Documents

    ELISA: Enzyme-linked Immunosorbent Assay PDF

    Description

    Learn about the competitive ELISA technique, a type of enzyme-linked immunosorbent assay used to detect and measure small proteins. Understand how it works and its application.

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