Immunology Quiz: Agglutination and Sensitization

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Questions and Answers

What is the primary purpose of Ficin in relation to RBCs?

To stabilize temperature during testing

To improve agglutination reactions

To enhance cell-cell contact

To cleave sialoglycoproteins from the RBC surface (correct)

At what temperature does IgM react best?

0°C to 4°C

30°C to 37°C

4°C to 27°C (correct)

37°C to 40°C

In passive (indirect) agglutination, what is typically used to detect the presence of antibodies?

Natural RBCs only

Antibodies directly from the serum

Soluble antigens alone

Insoluble particles coated with antigens (correct)

Which agglutination reaction requires the antigens to be present naturally on a particle?

Direct agglutination Signup and view all the answers

What is a key advantage of agglutination reactions?

They are quick and can be performed as needed Signup and view all the answers

What characterizes reverse passive agglutination?

Particles are coated with antibodies Signup and view all the answers

What is the ideal pH range for most antigen-antibody reactions to occur?

6.5-7.5 Signup and view all the answers

Which of the following tests uses known bacterial suspensions to check for antibodies in serum?

Widal test Signup and view all the answers

What is the primary purpose of the antihuman globulin-mediated agglutination technique?

To detect non-agglutinating antibodies using a second antibody. Signup and view all the answers

What is a direct antihuman globulin (Coombs') test primarily used to diagnose?

Autoimmune hemolytic anemia and hemolytic disease of the newborn. Signup and view all the answers

How does the indirect antihuman globulin (Coombs') test differ from the direct test?

It analyzes antibodies present in a patient sample through in vitro sensitization. Signup and view all the answers

What is the main reason for running positive and negative controls in agglutination tests?

To ensure the correct reagents are used and results are valid. Signup and view all the answers

What should be done if controls do not yield expected results during agglutination tests?

Discard the test as invalid and investigate possible errors. Signup and view all the answers

What is the main characteristic of agglutination?

It involves the aggregation of particles due to antigen-antibody reactions. Signup and view all the answers

Which of the following is NOT one of the types of particles used in agglutination reactions?

Viruses Signup and view all the answers

What occurs during the sensitization step of agglutination?

Antigen binds to antibody without cross-linking. Signup and view all the answers

What factor does NOT affect lattice formation in agglutination?

Time of incubation Signup and view all the answers

Why do erythrocytes repel each other in an ionic solution?

They carry a slight negative charge on their surface. Signup and view all the answers

Which enhancement technique reduces the surface charge of RBCs?

Using low ionic strength saline. Signup and view all the answers

What characteristic of IgM makes it a better agglutinin compared to IgG?

Larger molecular diameter and better bridging capacity. Signup and view all the answers

Which of the following statements about enhancement of lattice formation is false?

Enzymes can cleave groups to increase hydration of RBCs. Signup and view all the answers

What is the primary use of C-reactive protein (CRP) testing?

Detecting inflammation Signup and view all the answers

What principle does agglutination inhibition rely on?

Competition between soluble and particulate antigens Signup and view all the answers

What indicates a positive result in the agglutination inhibition test?

No agglutination is observed Signup and view all the answers

Which types of samples can be tested using the agglutination technique?

Wide variety of samples including blood and fecal samples Signup and view all the answers

What does a dark red, smooth button at the bottom of a microtiter well indicate?

Negative reaction Signup and view all the answers

Which antigen detection method uses red blood cells as the indicator particles?

Hemagglutination inhibition Signup and view all the answers

How is the titer determined in an agglutination test?

As the lowest sample concentration giving a positive result Signup and view all the answers

What is a potential application of the agglutination inhibition technique?

Detecting drugs such as heroin and cocaine Signup and view all the answers

Study Notes

Agglutination

Visible aggregation of particles caused by antigen-antibody reactions.
One of the reactants (Antigen or antibody) must be particulate.
Antibodies involved are called agglutinins.
The antigen must be exposed on the surface of the particle and each particle must have multiple epitopes.
Types of particles used: erythrocytes, bacterial cells, inert carriers like latex particles.

Sensitization

The initial reaction where the antigen binds to the antibody.
Each antibody molecule binds to a single epitope.
No cross-links or visible aggregates yet.
Affected by antibody affinity and epitope accessibility (exposed epitopes bind more readily).

Lattice Formation

Antibodies bridge the gaps between particles.
One antibody molecule binds to one epitope on each of two different particles.
Affected by:
- Ionic strength
- pH
- Temperature
- Ab:Ag ratio

Zeta Potential

Erythrocytes have a negative surface charge.
Cations surround cells in solution, forming an ionic cloud.
This charge repels cells, keeping them about 25 nm apart.
IgG antibodies can't bridge this distance due to size and hinge region flexibility.
IgM antibodies (35 nm diameter) are better agglutinins.

Enhancement Techniques

Reducing surface charges:
- Low ionic strength saline (LISS)
- Albumin (neutralizes surface charges)
- Enzymes (e.g., bromelin, papain, trypsin, ficin) cleave surface groups to decrease hydration
Centrifugation: Increases cell-cell contact.
Temperature:
- IgM: 4°C to 27°C (cold-reacting)
- IgG: 30°C to 37°C (warm-reacting)
pH: 6.5-7.5 is optimal for most Ag-Ab reactions.
Antihuman globulin reagent: Will be discussed later.

Advantages of Agglutination Reactions

Easy to perform.
Require simple equipment.
Relatively inexpensive.
Quick results.
Can be done as needed (no batching).
Generally qualitative, can be semi-quantitative.

Types of Agglutination Reactions

Direct: Naturally occurring antigens on particles (e.g., ABO blood grouping, Widal test).
Passive (indirect): Soluble antigen attached to an insoluble particle (e.g., detecting antibodies using antigen-coated particles).
Reverse Passive: Particles coated with antibody (e.g., detecting antigens using antibody-coated particles).
Agglutination Inhibition: Soluble antigens compete with particulate antigens for antibody binding sites (e.g., drug detection).

Direct Agglutination

Antigens present naturally on a particle.
Antibodies are added, resulting in visible agglutination.

Passive (Indirect) Agglutination

Soluble antigen is attached to a particle (RBCs, latex, gelatin).
Particles are coated with a known antigen to detect antibodies.

Reverse Passive Agglutination

Antibody-coated particles used to detect antigens.
Antibody is oriented for activity, with antigen-binding sites facing outward.

Agglutination Inhibition

Soluble antigens from the patient sample compete with indicator particles for antibody binding.
If the patient sample contains the target antigen, no agglutination occurs.
If the patient sample does not contain the target antigen, agglutination occurs.

Interpretation of Results

Semi-quantitative results obtained using serial dilutions.
Titer is the reciprocal of the highest dilution showing a positive result.
Performed in test tubes, microtiter plates, or on slides.

Tube Method

Tubes are centrifuged and shaken to see if the cell pellet can be evenly resuspended.

Microtiter Plate Method

Interpretation is based on cell sedimentation pattern.
Negative: Dark red, smooth button at the bottom.
Positive: Cells spread across the well's bottom.

Slide Method

Visual interpretation of latex or hemagglutination results.

Antihuman Globulin-Mediated Agglutination (Coombs' Test)

Detects reactions of non-agglutinating antibodies (IgG) by adding a second antibody.
Used when RBCs are sensitized with IgG but no visible agglutination occurs.
Antihuman globulin antibody recognizes and binds to the Fc region of human IgG bound to RBCs.
Bridging action allows lattice formation and agglutination.

Direct Antihuman Globulin Test

Detects in vivo sensitization of RBCs.
Used for diagnosing autoimmune diseases (e.g., autoimmune hemolytic anemia, hemolytic disease of the newborn).
Patient's RBC directly tested as they come from the body.

Indirect Antihuman Globulin Test

Determines the presence of a specific antibody in a patient sample.
Sensitization of RBCs occurs in vitro.

Agglutination: Quality Assurance

Run positive and negative controls for every test.
Follow manufacturer's instructions.
Store reagents properly.
Do not use expired reagents.

Summary

Agglutination reactions are essential for detecting antigens and antibodies.
Various techniques are used based on the type of antigen and antibody being tested.
Quality control measures are critical to ensure reliable results in all agglutination testing.

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Description

Test your knowledge on agglutination, sensitization, lattice formation, and zeta potential in immunology. Understand the interactions between antibodies and antigens and how these concepts contribute to immune responses. This quiz is essential for students of immunology and related fields.