Blood group serologyBlood group serology

The Nature of BloodThe Nature of Blood

The term The term blood blood refers to a highly refers to a highly complex mixture of cells, enzymes, complex mixture of cells, enzymes, proteins, and inorganic substances.proteins, and inorganic substances.

The fluid portion of blood is called The fluid portion of blood is called plasma. plasma. It is composed primarily of wIt is composed primarily of w

ater and accounts for 55% of blood ater and accounts for 55% of blood content.content.

Blood Solids:Blood Solids: Suspended in the plasma are these solid Suspended in the plasma are these solid

(cells) materials:(cells) materials: Red blood cells or erythrocytesRed blood cells or erythrocytes White blood cells or leukocytesWhite blood cells or leukocytes PlateletsPlatelets

SerumSerum The liquid that separates from the blood The liquid that separates from the blood

when a clot is formed. Blood clots when when a clot is formed. Blood clots when fibrin fibrin (a protein) traps and enmeshes the red (a protein) traps and enmeshes the red blood cells.blood cells.

Blood serology includes the study Blood serology includes the study of:of:

Red Blood Cells Antigens.Red Blood Cells Antigens. White Blood Cells Antigens.White Blood Cells Antigens. Platelet Antigens.Platelet Antigens. Antibodies in the serum.Antibodies in the serum. Compatibility studies for Compatibility studies for

transfusion purposes.transfusion purposes.

Antigen:Antigen:

A substance, foreign to the body that stimulates the A substance, foreign to the body that stimulates the production of antibodies by the immune system. production of antibodies by the immune system.

Were originally defined as, non-self molecules which Were originally defined as, non-self molecules which bound specifically to antibodies.bound specifically to antibodies.

Antigens which induce adaptive immunity are called Antigens which induce adaptive immunity are called immunogens. immunogens.

All immunogens are antigens, and are usually called All immunogens are antigens, and are usually called antigens unless their ability to induce an immune antigens unless their ability to induce an immune response is being changed. response is being changed.

Some antigens, called haptens, are not immunogenic Some antigens, called haptens, are not immunogenic unless they are covalently linked to immunogenic unless they are covalently linked to immunogenic carriers (usually proteins).carriers (usually proteins).

Hapten:Hapten: Small foreign molecule that is not antigenic. Small foreign molecule that is not antigenic. Must be coupled to a carrier molecule to be antigenic. Must be coupled to a carrier molecule to be antigenic. Once antibodies are formed they will recognize Once antibodies are formed they will recognize hapten.hapten.

The antigen bound by a unique antibody in a specific The antigen bound by a unique antibody in a specific

part which is called an epitope. Most proteins have part which is called an epitope. Most proteins have several epitopes that are recognized by different B several epitopes that are recognized by different B cells and induce a polyclonal antibody response.cells and induce a polyclonal antibody response.

Most are proteins or large polysaccharides from a Most are proteins or large polysaccharides from a foreign organism.foreign organism.

Microbes: Capsules, cell walls, toxins, viral Microbes: Capsules, cell walls, toxins, viral capsids, flagella, etc.capsids, flagella, etc.

Non-microbes: Pollen, egg white, red blood Non-microbes: Pollen, egg white, red blood cell surface molecules, serum proteins, and cell surface molecules, serum proteins, and surface molecules from transplanted tissue. surface molecules from transplanted tissue.

Lipids and nucleic acids are only antigenic Lipids and nucleic acids are only antigenic when combined with proteins or when combined with proteins or polysaccharides.polysaccharides.

Molecular weight of 10,000 or higher.Molecular weight of 10,000 or higher.

Epitope: Epitope:

Small part of an antigen that interacts with an Small part of an antigen that interacts with an antibody.antibody.

Any given antigen may have several epitopes.Any given antigen may have several epitopes.

Each epitope is recognized by a different Each epitope is recognized by a different antibody. antibody.

There are two types of immune response: There are two types of immune response: Negative Ag response:Negative Ag response:

Once Ag in the body, there is no response either Once Ag in the body, there is no response either humeral or cellular immunity " non responsive''. humeral or cellular immunity " non responsive''. e.g. if a e.g. if a patient accepted a given kidney. patient accepted a given kidney.

Positive Ag response.Positive Ag response.

AntibodyAntibody

A protein (immunoglobulin) produced by a B-cell A protein (immunoglobulin) produced by a B-cell that binds to a specific foreign antigen in the that binds to a specific foreign antigen in the blood or body fluids.blood or body fluids.

This leads to attack by the immune system. This leads to attack by the immune system. Belong to a group of serum proteins called Belong to a group of serum proteins called immunoglobulins (Igs).immunoglobulins (Igs).

Proteins that recognize and bind to a Proteins that recognize and bind to a particular antigen with very high specificity.particular antigen with very high specificity.

Made in response to exposure to the antigen.Made in response to exposure to the antigen.

One virus or microbe may have several One virus or microbe may have several antigenic determinant sites, to which different antigenic determinant sites, to which different antibodies may bind.antibodies may bind.

Each antibody has at least two identical sites Each antibody has at least two identical sites that bind antigen: that bind antigen: Antigen binding sites.Antigen binding sites.

Types of Ab:Types of Ab:

Allo Abs: Allo Abs:

Produced when the Ag is from the same species, e.g. Produced when the Ag is from the same species, e.g. from human to human.from human to human.

Auto Ab: Auto Ab:

Ag is a self Ag; stimulate production of Auto-Ab. Ag is a self Ag; stimulate production of Auto-Ab. e.g. immune haemolytic anaemias.e.g. immune haemolytic anaemias.

Hetero Ab:Hetero Ab:

Ag from a different species, e.g. from a camel to Ag from a different species, e.g. from a camel to human human

Brief Introduction to Antibody StructureBrief Introduction to Antibody Structure

The Ab takes a flexible Y-shaped molecule; it consists The Ab takes a flexible Y-shaped molecule; it consists of four polypeptides chains linked by disulphide of four polypeptides chains linked by disulphide bonds:bonds: 2 identical light chains "L" which is called kappa.2 identical light chains "L" which is called kappa. 2 identical heavy chains "H" which is called 2 identical heavy chains "H" which is called

lambda.lambda. Variable Regions: Two sections at the end of Y’s Variable Regions: Two sections at the end of Y’s

arms. Contain the antigen binding sites (Fab). arms. Contain the antigen binding sites (Fab). Identical on the same antibody, but vary from one Identical on the same antibody, but vary from one antibody to another.antibody to another.

Constant Regions: Stem of monomer and lower parts Constant Regions: Stem of monomer and lower parts of Y arms.of Y arms.

Fc region: Stem of monomer only. Important because Fc region: Stem of monomer only. Important because they can bind to complement or cells.they can bind to complement or cells.

Heavy chains have five different Heavy chains have five different isotypes which divide the Igs into isotypes which divide the Igs into five different classes, each with five different classes, each with different effector functions (in different effector functions (in humans IgG1-4, IgA1-2, IgD, humans IgG1-4, IgA1-2, IgD, IgM, IgE).IgM, IgE).

Ab classes:Ab classes:

Antibodies are grouped into one of five immunoglobulin Antibodies are grouped into one of five immunoglobulin classes:classes:

1- IgA (IgA1-2): alpha1- IgA (IgA1-2): alpha Structure: Dimer Percentage serum antibodies: 10-15% Location: Secretions (tears, saliva, intestine, milk), blood and lymph. Half-life in serum: 6 days Complement Fixation: No Placental Transfer: No Known Functions: Localized protection of mucosal surfaces.

Provides immunity to infant digestive tract. Called as secretory IgA (sIgA).

IgG (IgG1-4): Gamma. IgG (IgG1-4): Gamma. The most common serum antibody. It can pass from

mother to fetus during pregnancy, and diffuse across mucosal surfaces, the molecular weight is 160.000 D. IgG is also present in vaginal secretions, in higher concentrations than sIgA.

Structure: Monomer Percentage serum antibodies: 80% Location: Blood, lymph, intestine Half-life in serum: 23 days Complement Fixation: Yes Placental Transfer: Yes Known Functions: Enhances phagocytosis, neutralizes

toxins and viruses, protects fetus and newborn.

IgM: Mew. IgM: Mew.

Heavy molecular weight 1000.000. Structure: Pentamer Percentage serum antibodies:

5-10% Location: Blood, lymph, B cell surface (monomer) Half-life in serum: 5 days Complement Fixation: Yes Placental Transfer: No Known Functions: First antibodies produced during

an infection, before IgG. Effective against microbes and agglutinating antigens.

IgD: DeltaIgD: Delta Structure: Monomer Structure: Monomer

Percentage serum antibodies: 0.2% Percentage serum antibodies: 0.2% Location: B-cell surface, blood, and lymphLocation: B-cell surface, blood, and lymph Half-life in serum: 3 daysHalf-life in serum: 3 days Complement Fixation: NoComplement Fixation: No Placental Transfer: NoPlacental Transfer: No Known Functions: In serum function is unknown. Known Functions: In serum function is unknown.

On B cell surface, initiate immune response.On B cell surface, initiate immune response.

IgE: Epsilon. IgE: Epsilon. Structure: Monomer Percentage serum antibodies: 0.002% Location: Bound to mast cells and basophils

throughout body. Blood. Half-life in serum: 2 days Complement Fixation: No Placental Transfer: No Known Functions: Allergic reactions, possibly lysis

of worms.

Antigen-Antibody ReactionAntigen-Antibody ReactionThe interaction between the Ag and its specific Ab The interaction between the Ag and its specific Ab

to form a complex is mediated by multiple to form a complex is mediated by multiple bonds between the ADS and the ABS. these bonds between the ADS and the ABS. these bonds include:bonds include:

Hydrogen bonding.Hydrogen bonding. Electrostatic bonds.Electrostatic bonds. Vander waals bonding.Vander waals bonding. Hydrophobic bonding. Hydrophobic bonding.

-Hydrogen bonding results from formation of hydrogen bridges between appropriate atoms.

- The electrostatic forces results from attraction between oppositely charged groups.

- Vander Waals bonds generated by the interaction between electron clouds.

- The Hydrophobic bonds may contribute to about half of the total attractive forces between the Ag and Ab.

These bonds operate over a short distance; therefore the ADS must closely approach to ABS before interaction can occur.

Notes: Notes:

The Ag-Ab reaction is enhanced in presence The Ag-Ab reaction is enhanced in presence of electrolytes. of electrolytes.

The Ag-Ab reactions are characterized by:The Ag-Ab reactions are characterized by:

1) Being highly specific. 1) Being highly specific.

2) Depending in the non-covalent bonds; 2) Depending in the non-covalent bonds; therefore they are mostly reversibletherefore they are mostly reversible..

Consequences of Antigen-Antibody Consequences of Antigen-Antibody Binding:Binding:

Antigen-Antibody Complex: Formed when an antibody Antigen-Antibody Complex: Formed when an antibody binds to an antigen that it recognizes.binds to an antigen that it recognizes.

Affinity: A measure of binding strength.Affinity: A measure of binding strength.

1. Agglutination: 1. Agglutination:

2. Opsonization: 2. Opsonization:

3. Neutralization: 3. Neutralization:

4. Antibody-dependent cell-mediated cyto4. Antibody-dependent cell-mediated cytotoxicity: toxicity:

5. Complement Activation: 5. Complement Activation:

1. 1. Agglutination:Agglutination:

Antibodies cause antigens (microbes) to Antibodies cause antigens (microbes) to clump together.clump together.

IgM is more effective that IgG.IgM is more effective that IgG. Hemagglutination: Agglutination of red Hemagglutination: Agglutination of red

blood cells. Used to determine ABO blood cells. Used to determine ABO blood types and to detect influenza and blood types and to detect influenza and measles viruses. measles viruses.

2. Opsonization: 2. Opsonization: Antigen (microbe) is covered with antibodies Antigen (microbe) is covered with antibodies

that enhance its ingestion and lysis by that enhance its ingestion and lysis by phagocytic cells.phagocytic cells.

3. 3. Neutralization: IgG inactivates viruses by binding to their surface and

neutralize toxins by blocking their active sites.

4. Antibody-dependent cell-mediated cytotoxicity: Used to destroy large organisms (e.g.: worms). Target

organism is coated with antibodies and bombarded with chemicals from nonspecific immune cells.

5.Complement Activation: Both IgG and IgM trigger the complement system which

results in cell lysis and inflammation.

Erythroblastes Erythroblastes acidophilesacidophiles

DescriptionDescription• Hématies nuclééesHématies nucléées

ObservationObservation• Moelle stimulée ou irritéeMoelle stimulée ou irritée• Hyperrégénération de la Hyperrégénération de la

lignée rouge après lignée rouge après hémorragie ou hémolysehémorragie ou hémolyse

• DysérythropoièseDysérythropoièse• Infiltration maligneInfiltration maligne• MyélofibroseMyélofibrose• Post splénectomiePost splénectomie

Précipités Précipités d’Hémoglobine Hd’Hémoglobine H

DescriptionDescription• Granules très nombreux de Granules très nombreux de

petite taille visibles au au petite taille visibles au au bleu de crésylbleu de crésyl

ObservationObservation• Hémoglobinose HHémoglobinose H• Syndrome myéloprolifératifsSyndrome myéloprolifératifs• dysérythopoièsedysérythopoièse

Corps de HeinzCorps de Heinz DescriptionDescription

• Corpuscules de 0,3µm de Corpuscules de 0,3µm de diamètre visibles après coloration diamètre visibles après coloration spéciale par l e bleu de crésyl spéciale par l e bleu de crésyl brillant correspondant à brillant correspondant à l’hémoglobine dénaturéel’hémoglobine dénaturée

ObservationObservation• Hémolyse toxiqueHémolyse toxique• Hémolyse par des agents Hémolyse par des agents

oxydants chez les sujets déficients oxydants chez les sujets déficients en glutathion ou ayant une en glutathion ou ayant une Hémoglobine instableHémoglobine instable

• ß ß TthalassémieTthalassémie• Post splénectomiePost splénectomie

• MGG d’après atlas hématologie F.G.H MGG d’après atlas hématologie F.G.H Hayhoe editions Flammarion 1970Hayhoe editions Flammarion 1970

ParasitesParasites

Plasmodium falciparumPlasmodium falciparum

Plasmodium vivaxPlasmodium vivax

• Lames du service Lames du service d’hématologie de l’hôpital de d’hématologie de l’hôpital de Saint Denis (93) 2002Saint Denis (93) 2002

Plasmodium falciparumPlasmodium falciparum

PhotosPhotos Microscopie Electronique Microscopie Electronique

D’après l’ouvrageD’après l’ouvrage

« les Cellules du Sang »« les Cellules du Sang »

M.BESSISM.BESSIS

Editions Masson 1972 avec leur autorisationEditions Masson 1972 avec leur autorisation Microscopie optiqueMicroscopie optique

Viviane GUILLAUMEViviane GUILLAUMELames provenant pour la plupart du service d’ hématologieLames provenant pour la plupart du service d’ hématologie

CHU La Pitié Salpêtrière pendant la préparation du DERBHCHU La Pitié Salpêtrière pendant la préparation du DERBH

et du CHU de Nancy avec l’aimable autorisation de Mr le et du CHU de Nancy avec l’aimable autorisation de Mr le DrVISANICADrVISANICA