Podcast
Questions and Answers
What is the primary advantage of indirect immunofluorescence over direct immunofluorescence?
What is the primary advantage of indirect immunofluorescence over direct immunofluorescence?
In the context of the immunochromatography test, what is the role of the control line?
In the context of the immunochromatography test, what is the role of the control line?
Which component is added last in the ELISA method described?
Which component is added last in the ELISA method described?
What method uses dye-labelled antibodies to detect antigens in serum or microorganisms?
What method uses dye-labelled antibodies to detect antigens in serum or microorganisms?
Signup and view all the answers
Which process results in the formation of a precipitate from the interaction between antibodies and antigens?
Which process results in the formation of a precipitate from the interaction between antibodies and antigens?
Signup and view all the answers
Which type of immunofluorescence directly tags the antibody specific to the antigen?
Which type of immunofluorescence directly tags the antibody specific to the antigen?
Signup and view all the answers
What is the primary purpose of the labeled antibody in immunochromatography?
What is the primary purpose of the labeled antibody in immunochromatography?
Signup and view all the answers
Which immunological test is used for the diagnosis of syphilis?
Which immunological test is used for the diagnosis of syphilis?
Signup and view all the answers
What is the main principle behind direct agglutination reactions?
What is the main principle behind direct agglutination reactions?
Signup and view all the answers
Which test uses agglutination principles to diagnose syphilis?
Which test uses agglutination principles to diagnose syphilis?
Signup and view all the answers
In indirect agglutination tests, what surface is NOT commonly used to bind antigens?
In indirect agglutination tests, what surface is NOT commonly used to bind antigens?
Signup and view all the answers
What does the presence of clumping in a slide test indicate?
What does the presence of clumping in a slide test indicate?
Signup and view all the answers
Which agglutination reaction is primarily used for blood grouping?
Which agglutination reaction is primarily used for blood grouping?
Signup and view all the answers
Which of the following is a characteristic of agglutination inhibition tests?
Which of the following is a characteristic of agglutination inhibition tests?
Signup and view all the answers
The hemagglutination test detects interactions between what two components?
The hemagglutination test detects interactions between what two components?
Signup and view all the answers
Which of the following is NOT an application of indirect agglutination tests?
Which of the following is NOT an application of indirect agglutination tests?
Signup and view all the answers
What does the term 'titer' refer to in serological reactions?
What does the term 'titer' refer to in serological reactions?
Signup and view all the answers
Which of the following is NOT a type of serological reaction?
Which of the following is NOT a type of serological reaction?
Signup and view all the answers
What is the primary disadvantage of the direct ELISA method?
What is the primary disadvantage of the direct ELISA method?
Signup and view all the answers
In the indirect ELISA method, what is the role of the labeled secondary antibody?
In the indirect ELISA method, what is the role of the labeled secondary antibody?
Signup and view all the answers
Which enzyme is commonly used in ELISA techniques?
Which enzyme is commonly used in ELISA techniques?
Signup and view all the answers
What type of serological reaction uses a known antibody to detect an unknown antigen?
What type of serological reaction uses a known antibody to detect an unknown antigen?
Signup and view all the answers
What is a common application of ELISA in disease management?
What is a common application of ELISA in disease management?
Signup and view all the answers
What is one of the substrates used in the detection process of ELISA?
What is one of the substrates used in the detection process of ELISA?
Signup and view all the answers
Study Notes
General Microbiology and Immunology, PMB201, Lecture 4B
- Serology is a branch of laboratory medicine that tests for infection.
- It evaluates antigen-antibody reactions in vitro.
- It can detect and identify unknown antibodies using known antigens.
- It can detect and identify unknown antigens using known antibodies.
- Titer refers to the highest dilution of antibody that produces a visible reaction with an antigen.
Types of Serological Reactions
- ELISA (Enzyme-linked immunosorbent assay)
- Precipitation
- Agglutination
- Neutralization test
- Complement fixation test (CFT)
- Immunofluorescence (IF)
- Radioimmunoassay (RIA)
- Blotting techniques
Enzyme-linked immunosorbent assay (ELISA)
- It is an immunoassay technique that involves the reaction of antigen and antibody in vitro.
- ELISA is a sensitive and specific assay for the detection (and quantitation) of antigens or antibodies.
- Principle: The antigen is immobilized to a solid surface(directly or by a capture antibody).
- The antigen is detected by an antibody conjugated with an enzyme (horseradish peroxidase (HRP) or alkaline phosphatase (AP)).
- The enzyme is detected by a substrate (e.g., 3,3',5,5'-Tetramethylbenzidine (TMB) or p-Nitrophenyl Phosphate (PNPP)).
Applications of ELISA
- Serum antibody concentrations
- Disease outbreaks tracking
- Detection of antibodies in blood samples for previous infection.
- Detection of antigens (e.g., pregnancy hormones, drug allergens)
- Detection of potential food allergens (e.g., milk, peanuts, walnuts, almonds, and eggs)
Types of ELISA
- Direct ELISA
- Indirect ELISA
- Sandwich ELISA
- Competitive ELISA
Direct ELISA
- Uses a labelled primary antibody that directly reacts with the antigen.
- Disadvantages: Immunoreactivity of the primary antibody is affected by labeling with enzymes or tags.
Indirect ELISA
- Antigen is bound to a solid phase
- Antibody tested serum (primary antibody) is added
- Known labeled secondary antibody binds with enzyme.
- Measure enzyme label
Sandwich ELISA
- Plate is coated with a capture antibody
- Sample is added and antigen binds to the capture antibody
- Primary antibody binds to the antigen
- Enzyme-linked secondary antibody binds to the detecting antibody.
- Substrate is added, converted into a detectable form by the enzyme
Immunofluorescence test
- Fluorescent dyes (e.g., fluorescein or rhodamine) are attached to known specific antibodies (Abs) to detect the presence of antigens (Ag) in serum or microorganisms in a sample.
- It's valuable for identifying and locating antigens on cell surfaces or tissues.
- Types: Direct and Indirect
Direct Immunofluorescence
- The antibody specific to the antigen is directly tagged with the fluorochrome.
- Used in detecting rabies virus in brain smears.
Indirect Immunofluorescence
- The antibody specific for the antigen is unlabeled.
- A second antibody directed toward the first antibody is tagged with the fluorochrome.
- Indirect fluorescence is more sensitive due to signal amplification.
- Used in detecting syphilis.
Immunochromatography (Rapid test)
- Speed method for the etiologic diagnosis of infections
- Dye-labelled antibody specific for the target antigen is present on the end of a nitrocellulose strip/plastic well
- Antibody specific for the target antigen binds to the strip
- Either antibody specific for the labelled antibody or the antigen is bound at the control line
- Test (T): Presence/absence of Antigen or Antibody determined.
- Control (C): Assesses Validity of the Test.
- Applications: HCV (serum antibody), HBC (serum antigen), pregnancy hormones (HCG)
Precipitation reactions
- Interaction between antigen and antibody molecules resulting in precipitate formation from solution.
- Types: Ring test, Tube test.
Ring test
- Ascoli's thermoprecipitin test, used for diagnosing anthrax.
Tube test
- A simple quantitative tube precipitation test (e.g., Kahn's test) for syphilis.
Slide test
- Antigen and antibody mixed on a slide producing floccules (for positive tests), used for VDRL syphilis test.
Immunodiffusion
- Elk test for toxigenicity in diphtheria bacilli
Agglutination reactions
- Interaction between antibody and antigen resulting in visible clumping (agglutination).
- If the antigen is an erythrocyte, the interaction is called hemagglutination.
- Types: Direct agglutination tests, Indirect agglutination tests, Hemagglutination inhibition.
Direct Agglutination Reaction
- Detects soluble antibodies in serum using specific antigen.
- Serum is serially diluted.
- Antigen is added, mixed, and incubated.
- Results are expressed as a titer (highest dilution with positive agglutination).
- Applications: blood grouping, brucellosis, typhoid (enteric fever).
Widal Test
- Rapid slide test or quantitative tube test
- Detects presence/absence of S. Typhi and S. Paratyphi antibodies
- Results interpreted based on antibody rise, signifying recent or convalescent stage infection
Indirect Agglutination
- Tests for antibodies in patient serum using antigens bound to artificial surfaces (latex, RBCs, bentonite, or charcoal).
- Antibody reaction to antigens leads to agglutination of carrier particles (visible clumps).
- Applications: pregnancy hormone assays, rheumatoid arthritis diagnosis (Rose-Waaler test)
Agglutination Inhibition
- Tests for soluble antigens in patient serum using reagent antibody specific for the antigen and reagent antigen (latex or RBCs).
- Presence of antigen blocks reagent antibody from binding to particles, thereby inhibiting agglutination and showing absence antigen.
- Applications: virus infection detection, illegal drug detection.
The Adaptive or Specific Immune System
- It takes effects after the innate immune response.
- It's the third line of defense against infection.
- It is specific, acquired, and has immune memory.
- It involves B and T lymphocytes
Adaptive Immunity: Functions
- Recognizes specific antigens.
- Eliminates specific pathogens or infected cells.
- Develops immunological memory (memory cells).
Adaptive Immunity: Composition
- Cellular immunity (T-cells):
- Eliminates pathogens that have invaded cells (intracellular infection).
- Humoral immunity (B-cells):
- Provides antibodies to bind pathogens and their products (extracellular infection).
Cell Surface Molecules: Major Histocompatibility Complex (MHC)
- Each cell possesses glycoprotein surface molecules (MHC or HLA).
- Class I MHC: found on the surface of all nucleated cells; CD8+ cytotoxic T cells recognize antigens associated with MHC class I.
- Class II MHC: found on the surface of Antigen-presenting cells; CD4+ Helper T cells recognize antigens in association with MHC class II.
T Cell-Mediated Immune Response: Antigen Recognition
- T cells develop in bone marrow and mature in the thymus.
- Naïve T cells move to secondary lymphoid organs.
- Naïve T cells recognize antigens presented by antigen-presenting cells (e.g., dendritic cells, macrophages, B cells).
T Cell Activation
- Signal 1: TCR binds to antigen-MHC combination.
- Signal 2: Costimulatory molecules (e.g., CD28) bind, activating the T cell.
- Signal 3: Cytokines from antigen-presenting cells activate the T cell to an effector phenotype.
Anergy
- State of immune unresponsiveness.
- Induced when the TCR is stimulated without a co-stimulatory signal.
T Cell Proliferation and Differentiation
- Naive T cells respond to different antigens presented through MHC.
- Naive T cells proliferate into effector T cells.
- Effector T cell types: Helper T cells (Th1, Th2, Th17, Tfh), Cytotoxic T cells (Tc), Regulatory T cells (Treg), Memory T cells.
T-Effector Cells (Helper T Cells)
- Do not directly kill invading pathogens.
- Have no phagocytic or cytotoxic activity.
- Help other immune cells (e.g., by activating the immune response and releasing cytokines).
- Helper T cells express CD4 receptors that bind peptides displayed by class II MHC molecules.
Exogenous Antigens
- Inhaled, ingested, or injected antigens are taken up by antigen-presenting cells (APCs), such as dendritic cells and macrophages.
- Antigens are degraded into fragments (short peptides).
- The fragments are displayed with MHC class II molecules.
- Helper T (CD4+/Th) cells recognize the antigen.
- CD4+/Th cells release cytokines to stimulate the humoral immune response and antibody production.
###Humoral Immunity: B Cells
- Produced in bone marrow.
- Involved in humoral immunity.
- Migrate to secondary lymphoid tissues (lymph nodes, spleen, lymphatic tissue).
- Activated into plasma cells and memory cells.
B Cell Activation
- Activation mechanisms: T-dependent antigen activation, T-independent antigen activation.
T-Dependent Antigens
- Requires Th cell help.
- Most antigens are proteins
- Activation of B cells by T cells results in plasma and memory cells.
T-Independent Antigens
- Direct activation of B cells by antigens.
- Antigens (e.g., polysaccharides) can cross-link B cell receptors and induce activation without T-cell help.
- Plasma cells are activated, but no memory cells are generated.
Studying That Suits You
Use AI to generate personalized quizzes and flashcards to suit your learning preferences.
Related Documents
Description
This quiz covers essential concepts from Lecture 4B of PMB201, focusing on serology and its various applications in laboratory medicine. Key topics include the evaluation of antigen-antibody reactions, types of serological reactions, and the principles of the ELISA technique. Test your understanding of these critical immunology topics!
