Agglutination Reactions

December 9, 2013

Agglutination
RBC

IgM Antibody

RBC

RBC

RBC RBC RBC RBC

RBC IgG Antibody RBC RBC RBC

Agglutination Reactions

Characteristics
is the aggregation of particulate matter due to combination with specific antibody. takes place on the surface of the particle

and thus antigen must be exposed and able

to bind with antibody

Involve particulate antigens and antibodies

Characteristics contd
types of particles participating in such reactions include:

erythrocytes, bacterial cells, and inert carrier such as

latex particles. antigen consists of suspensions of microorganisms, cells (rbc) or uniform particles like latex Antigens may be:
On a cell (direct agglutination) Attached to latex spheres (indirect or passive agglutination)

Characteristics contd
Agglutination reaction is aided by elevated

temperature (37-56C) and by:

Movement which increases the contact between antigen and antibody. Clear supernatant. Clumps aggregate and settle as large visible clumps.

Agglutination Requirements
Availability of stable cell or particle suspension Presence of one or more antigens close to

the surface
Knowledge that incomplete or non-

agglutinating antibodies are not detectable

without modifications, e.g. antiglobulin (Coombs) technic

Advantages
Agglutination of insoluble native antigens or antigen-coated particles simple to read with or without the aid of a microscope.

Increased degree of sensitivity

Great variety of detectable substances

Steps in Agglutination
1. Sensitization

involves antigen-antibody combination through

single antigenic determinants on the particle surface

may be affected by the nature of the antibody molecules, affinity and avidity of individual

antibody, and the nature of antigen-bearing

surface

Steps in Agglutination Reaction

2. Aggregative Stage represents the sum of interaction between antibody and multiple antigenic determinants on a particle dependent on environmental conditions as well as the

relative concentrations of antigen and antibody

it could be enhanced by LISS (Low ionic strength saline), neutralizes surface charges and facilitates agglutination

(albumin 5 - 50%)

Uses of Agglutination Reactions

Aid in the identification, by means of known antisera (serum containing antibodies speciftic for a given antigen), microorganisms cultured from clinical

specimens.

Help estimate the titer of antibacterial agglutinins in the serum of patients with unknown diseases.

Types of Agglutination Reactions

Direct Agglutination Passive Agglutination Reverse Passive Agglutination Agglutination inhibition Hemagglutination-inhibition Coagglutination/Conglutination

Types of Agglutination Rxns.

Direct agglutination - occurs when antigens are found naturally on a particle (Serotyping of Salmonella)

e.g. identification of bacterial types

O antigen (somatic) - compact, fine and granular agglutination H antigen (flagellar) - form a loosely woven network of clumped cells (coarse and floccular), called snowflake

agglutination
hemagglutination (agglutination reaction which involves rbc)

Direct Agglutination Reaction

Agglutination Reactions
Passive agglutination - employs particles that are coated with antigens not normally found on their surfaces. Inert materials commonly used: 1. Bentonite 2. Latex particles 3. Colloidion 4. Colloidal charcoal

Agglutination Reactions
Reverse Passive agglutination - antibody rather than

antigen is attached to a carrier particle.

Agglutination inhibition - based on competition between particulate and soluble antigens for limited antibody combining sites, and a lack of agglutination is an indicator of a positive reaction. Hemagglutination inhibition reactions -tests the presence of antibodies to certain viruses, such as rubella, mumps, measles, influenza, and adenovirus

Hemagglutination
Hemagglutination involves agglutination of RBCs. Viral hemagglutination inhibition tests for antibodies by the

antibodies' ability to prevent viruses from agglutinating RBCs.

Figure 18.7

Neutralization Reactions
Eliminate the harmful effect of a virus or exotoxin

Figure 18.8b

Agglutination Reactions
Coagglutination/Conglutination - name given to

systems using bacteria as inert particles to which

antibody is attached (S. aureus).
the Fc region of antibody attaches to protein A of staphylococcal cell leaving the Fab region to combine with the antigen killed staphylococcal cells coated with antibody can be used to identify bacteria and detect soluble extracellular bacterial antigens in specimens and body fluids.

Reading Agglutination Reactions

Macroscopic
+4 +3 One solid aggregate or clump of cells. Several large aggregates, clear background.

+2
+1

Small to medium sized aggregates, clear background.

Small aggregates, turbid reddish background. Weak granularity in cell suspension. A few macroscopic agglutinates but numerous agglutinates microscopically

MF

Any degree of agglutination in a sea of unagglutinated cells

Hem Hemolysis is interpreted as a positive reaction and may be graded as complete or partial; hemolysis and agglutination may be recorded on the same tube. No agglutination, smooth reddish background

Microscopic: + Positive - aggregates of at least 3-5 cells. Negative - no agglutination.

Agglutination Reactions

Microscopic Agglutination

Hemagglutination

Hemagglutination Inhibition

Agglutination Reactions

Latex Agglutination