Immunology Flashcards
Define an Antibody (Ab) ?
An immunoglobulin (Ig), is a large, Y-shaped protein produced mainly by B lymphocytes of the immune system
Define an Antigen (Ag) ?
A substance that induces an immune response e.g. antibodies
Define Epitope ?
Part of the antigen that the antibody attaches it self to
Define Immunoassay/assay ?
A procedure for detecting or measuring specific proteins or other substances through their properties as antigens or antibodies
Define Serology/serological assay ?
The scientific study or diagnostic examination of blood serum
Define Analyte ?
The molecule being measured in the assay
Define Enzyme-linked immunosorbent assay (ELISA) ?
A quantitative immunological assay used to measure proteins such as antibodies and antigens
Define Monoclonal antibodies ?
Antibodies that are made by identical immune cells that are all clones of a unique parent B cell
Define Polyclonal antibodies ?
Antibodies that are secreted by different B cell lineages within the body
What is the membrane bound for antibodies ?
Known as Immunoglobulin or B cell receptor (BCR)
What can the Variable Region and the constant region also be called ?
Variable Region= Fab region
Constant region= Fc region
What are examples of antigens ?
Can be pathogen components, proteins, carbohydrates, lipids, drugs, toxins…etc
Monoclonal vs. Polyclonal antibodies ?
- Highly specific for corresponding antigen, detect 1 epitope out of >108
- Easy to isolate & study. Invaluable as probes of biological processes
- Have different Isotypes (Ab class) conferring diverse functions
- Can be generated in large amounts
- Have high Affinity for their antigen
What does the antibody affinity constant affect ?
Sensitivity
What is avidity ?
The total strength of the interaction
How is the affinity measured ?
AFFINITY is measured by KD - equilibrium dissociation constant
ELISA has ?
High sensitivity, but not necessarily high specificity
What does ELISA diagnose?
Diagnosis of HIV infection, pregnancy tests, measurement of cytokines or soluble receptors in cell supernatant or serum
ELISA assays are generally carried out in ? allowing ?
ELISA assays are generally carried out in 96 well plates, allowing multiple samples to be measured in a single experiment.
Explain the Indirect ELISA practical ?
- Microwell plates are incubated with antigens, washed (to remove unbound) and blocked with BSA
- Samples and controls are added and incubated
- Plates are washed and then an enzyme-linked secondary antibody is added and plate washed
- Substrate is added and enzymes on the antibody elicit a colourometric change
What does a Positive control confirm ?
It confirms the assay is working and is specific
What does a Negative control confirm ?
It confirms a positive result is not due to non-specific binding of Abs or faulty reagents or any other reason
Each negative control has one vital step removed:
- NO antigen (BSA)
- NO primary antibody
- NO Labelled detection/secondary Ab
- NO Substrate
What is Analytical Sensitivity ?
– ability of a test to detect very small amounts of Ag or Ab
What is Clinical Sensitivity?
– ability of test to give positive result if patient has the disease (no false negative results)
Explain a False negative ?
– the test result is negative, as the concentration of the measured Ab or Ag is too low for detection (sensitivity). In reality, the patient has the disease and the test result should be positive
What is Analytical Specificity ?
– ability of test to detect substance without interference from cross-reacting substances
What is Clinical Specificity?
– ability of test to give a negative result if patient does not have disease (no false positive results)
Explain a False positive ?
– the test result is positive, as the test is detecting another cross-reacting substance. In reality, the test result should be negative as the patient does have the disease
