Immune SystemImmune System

Cells and Cell Products That Cells and Cell Products That Protect the Body Against Protect the Body Against DiseaseDisease

T cells B cells

Properties of the Immune ResponseProperties of the Immune Response

A.A. Specificity: Unique response Specificity: Unique response against each each invaderagainst each each invader

B. Memory: Increased response at B. Memory: Increased response at next invasionnext invasion

C. Discrimination: Ability to C. Discrimination: Ability to distinguish “self” from “non-self” distinguish “self” from “non-self”

Applying Your KnowledgeApplying Your Knowledge

Which property of the immune response: Which property of the immune response:

A.A. keeps you from developing chicken pox a keeps you from developing chicken pox a second time? (Assume you had a severe second time? (Assume you had a severe case at age six.)case at age six.)

B.B. helps you successfully recover from a cold?helps you successfully recover from a cold?

C.C. causes you to reject a donated organ?causes you to reject a donated organ?

1.1. SpecificitySpecificity2.2. MemoryMemory3.3. Discrimination between Discrimination between

Self and Non-selfSelf and Non-self

Sources and Transport of Immune System CellsSources and Transport of Immune System Cells

• Sites of immune Sites of immune cell production or cell production or maturationmaturation

• bone marrow bone marrow • spleenspleen• thymusthymus• tonsilstonsils

• CirculationCirculation– lymphatic system lymphatic system

• lymph vesselslymph vessels• lymph nodeslymph nodes

– bloodstreambloodstream

Cells of the Cells of the Immune SystemImmune System

B cells produce antibodies that

bind to antigens on the surface of

the invader

T cells carry out cell-cell

interactions

Macrophages engulf invaders

and provide signals for other immune system

cells

Activities of Activities of MacrophagesMacrophages

• Surround invader Surround invader with plasma with plasma membrane and membrane and bring it into cellbring it into cell(phagocytosis)(phagocytosis)

• Display antigen on Display antigen on surface by making surface by making a complex with a complex with MHC proteinMHC protein

MHC= major histocompatibility complex, MHC= major histocompatibility complex, 70 genes that code for cell surface proteins70 genes that code for cell surface proteins

B Cells Fight Against B Cells Fight Against Viruses and BacteriaViruses and Bacteria

• Each B cell carries a unique antibody on its Each B cell carries a unique antibody on its surfacesurface

• When antigen on surface of invader binds to When antigen on surface of invader binds to antibody on B cell, the B cell divides and gives antibody on B cell, the B cell divides and gives rise to two types of cellsrise to two types of cells

--plasma cell: actively secretes antibody--plasma cell: actively secretes antibody into bloodstream into bloodstream

--memory B cell: responds on second --memory B cell: responds on second invasion invasion

Primary Response: initial

activation of immune system and destruction

of invader

Secondary Response: faster

and stronger activation of

immune system due to memory

cells

Memory Cells and the Immune Response

Antibody StructureAntibody Structure

Antibody has 2 heavy chains and 2 light chains.Antibody has 2 heavy chains and 2 light chains.Each chain has variable (V) and constant (C) region.Each chain has variable (V) and constant (C) region.Variable regions bind to antigen.Variable regions bind to antigen.Constant regions bind to cells or other antibodies.Constant regions bind to cells or other antibodies.

Gene forGene forconstant regionconstant regionof light chainof light chain

Genes forGenes forvariable regionvariable regionof light chainof light chainGene forGene for

constant regionconstant regionof heavy chainof heavy chain

Genes forGenes forvariable regionvariable regionof heavy chainof heavy chain

Recombination during Recombination during Construction of Antibody GenesConstruction of Antibody Genes

Antibodies Mark Invaders for DestructionAntibodies Mark Invaders for Destruction

Phagocytosis by Phagocytosis by a Macrophagea Macrophage

Applying Your KnowledgeApplying Your Knowledge

A.A. Which part of the antibody binds to the Which part of the antibody binds to the antigen? (choices on left)antigen? (choices on left)

B.B. Which type of protein structure is seen Which type of protein structure is seen in a functional antibody molecule? in a functional antibody molecule? (choices on right)(choices on right)

1.1. Light ChainLight Chain2.2. Heavy ChainHeavy Chain3.3. Variable RegionVariable Region4.4. Constant RegionConstant Region

1.1. Primary StructurePrimary Structure2.2. Secondary StructureSecondary Structure3.3. Tertiary StructureTertiary Structure4.4. Quaternary StructureQuaternary Structure

Classes of T CellsClasses of T Cells1.1. Helper T cells: activate B Helper T cells: activate B

cells and cytotoxic T cells cells and cytotoxic T cells

2.2. Cytotoxic T cells: rupture Cytotoxic T cells: rupture infected body cells or infected body cells or cancer cellscancer cells

T cell

Cancer cell

Cytotoxic T cell Cytotoxic T cell attacks a cancer cell attacks a cancer cell

Communication Communication

Among CellsAmong Cells

1. Macrophage engulfs invader

2. Macrophage displays

“processed” antigen

5. Macrophage releases cytokines to activate Helper T cell

4. Helper T Cell binds to

Macrophage 3. Invader binds to B cell that carries antibody matching

the antigen

6. Activated Helper T cell stimulates

division of selected B cell

7. Dividing B cell gives rise to

memory B cells and plasma cells

Applying Your KnowledgeApplying Your Knowledge

Which immune system cell: Which immune system cell:

A.A. keeps you from developing chicken pox a keeps you from developing chicken pox a second time? (Assume you had a severe second time? (Assume you had a severe case at age six.)case at age six.)

B.B. helps you successfully recover from a cold?helps you successfully recover from a cold?

C.C. causes you to reject a donated organ?causes you to reject a donated organ?

1.1. MacrophageMacrophage2.2. Plasma CellPlasma Cell3.3. Memory B CellMemory B Cell4.4. Helper T CellHelper T Cell5.5. Cytotoxic T CellCytotoxic T Cell

Humoral and Cellular ImmunityHumoral and Cellular Immunity

Humoral Humoral

(Antibody-mediated)(Antibody-mediated)

CellularCellular

(T-cell mediated)(T-cell mediated)

Hay feverHay fever Reaction to poison ivyReaction to poison ivy

Transfusion reactionsTransfusion reactions Tuberculin TestTuberculin Test

Rh diseaseRh disease Reaction to bee stingsReaction to bee stings

ImmunizationImmunization Graft (organ) rejectionGraft (organ) rejection

Viral and Bacterial Viral and Bacterial infectionsinfections

Cancer PreventionCancer Prevention

Diseases of the Immune Diseases of the Immune SystemSystem

A.A. Inherited Immune DeficienciesInherited Immune Deficiencies

eg. SCID: eg. SCID: Severe Combined Immune DeficiencySevere Combined Immune Deficiency

B. AIDS: B. AIDS: Acquired Immune Deficiency SyndromeAcquired Immune Deficiency Syndrome

C. Autoimmune diseasesC. Autoimmune diseases

1. Scleroderma1. Scleroderma

2. Rheumatiod arthritis2. Rheumatiod arthritis

D. Allergies D. Allergies

AllergiesAllergiesProduces

antibodies with unique constant

region (IgE)

IgE antibodies

bind to mast cells

Mast cells release

histaminesAntigen binds to IgE on

mast cells

Antigen binds to B

cell and activates it

HIVHIV

GlycoproteinsGlycoproteinsGlycoproteinsGlycoproteins

EnvelopeEnvelope(lipid bilayer)(lipid bilayer)

EnvelopeEnvelope(lipid bilayer)(lipid bilayer)

ProteinProteinCoatCoat

ProteinProteinCoatCoat

CoreCoreProteinsProteins

CoreCoreProteinsProteins

ReverseReverseTranscriptaseTranscriptase

ReverseReverseTranscriptaseTranscriptase

Viral RNA inViral RNA inprotein coatprotein coatViral RNA inViral RNA inprotein coatprotein coat

Entry of HIV Retrovirus into Host CellEntry of HIV Retrovirus into Host Cell

DNA copy of retroviral genome

Production of New Viral ParticlesProduction of New Viral Particles

Contains more than one protein product; individual proteins are separated by

protease

HIV Binds to CD4 and CCR5 HIV Binds to CD4 and CCR5 Co-receptors on Helper T cellsCo-receptors on Helper T cells

Individuals that are homozygous for a deletion Individuals that are homozygous for a deletion mutation in the CCR5 gene are resistant to HIV infection.mutation in the CCR5 gene are resistant to HIV infection.

HIV Destroys T cellsHIV Destroys T cells

Use of ELISA to detect Use of ELISA to detect Antibodies to HIVAntibodies to HIV

ELISA = Enzyme-Linked ELISA = Enzyme-Linked Immunosorbent Assay Immunosorbent Assay

Tests serum for the presence of Tests serum for the presence of antibodies directed against HIVantibodies directed against HIV

ELISAELISA ProcedureProcedure

HIV antigenHIV antigen HIV antigenHIV antigenInfected PersonInfected Person Uninfected PersonUninfected Person

No Anti-HIV in serumNo Anti-HIV in serumof uninfected personof uninfected person

Anti-HIV in serumAnti-HIV in serumof infected personof infected person

Anti-IgG with Anti-IgG with horseradish horseradish peroxidase enzymeperoxidase enzyme

Anti-IgG with enzyme Anti-IgG with enzyme does not binddoes not bind

Substrate:Substrate:SalicylateSalicylate(colorless)(colorless)

Product:Product:Oxidized Oxidized SalicylateSalicylate(brown(brown))

No brown product No brown product formedformed

ResultResult ResultResult

Instructions for ELISAInstructions for ELISA

• Follow ALL directions on pages 90-101 of Follow ALL directions on pages 90-101 of workbook (summarized in these slides)workbook (summarized in these slides)

• Label theLabel the vertical side of a vertical side of a microtiter platemicrotiter plate as as Rows 1, 2, 3, 4 so that there are three wells per Rows 1, 2, 3, 4 so that there are three wells per row. row.

• Add the antigenAdd the antigen: Use a micropipettor and a : Use a micropipettor and a CLEAN tip to add 0.1 mL (100 uL) of HIV antigen CLEAN tip to add 0.1 mL (100 uL) of HIV antigen (viral antigens) to all wells.(viral antigens) to all wells.

• IncubateIncubate at room temperature for 5 minutes then at room temperature for 5 minutes then empty the wells with the “flip and bang” method.empty the wells with the “flip and bang” method.

• Wash the wells:Wash the wells: Add PBS buffer with transfer Add PBS buffer with transfer pipette, empty wells with “flip and bang”  method. pipette, empty wells with “flip and bang”  method.

Instructions for ELISAInstructions for ELISA• Add the test sera:Add the test sera: (use clean tip for each) (use clean tip for each)

– 100 uL PBS buffer to the three wells in row 1 100 uL PBS buffer to the three wells in row 1 (negative control).(negative control).

– 100 uL of “+” (positive control: antibody to the 100 uL of “+” (positive control: antibody to the HIV antigen) to the three wells in row 2. HIV antigen) to the three wells in row 2.

– 100 uL of Donor Serum 1 to the three wells in 100 uL of Donor Serum 1 to the three wells in row 3.row 3.

– 100 uL of Donor Serum 2 to the three wells in 100 uL of Donor Serum 2 to the three wells in row 4.row 4.

• IncubateIncubate at 37 at 37oo (oven on side bench) for 15 (oven on side bench) for 15 minutes then minutes then empty the wellsempty the wells with the “flip and with the “flip and bang” method.bang” method.

• Wash the wells:Wash the wells: Add PBS buffer with transfer Add PBS buffer with transfer pipette, empty wells with “flip and bang”  method. pipette, empty wells with “flip and bang”  method.

Instructions for ELISAInstructions for ELISA

• Add the 2Add the 2oo Ab: Ab: Place 100 uL of the anti-IgG Place 100 uL of the anti-IgG peroxidase conjugate in all 12 wells.peroxidase conjugate in all 12 wells.

• IncubateIncubate at 37 at 37oo (oven on side bench) for 15 (oven on side bench) for 15 minutes then minutes then empty the wellsempty the wells with the “flip and with the “flip and bang” method.bang” method.

• Wash the wells:Wash the wells: Add PBS buffer with transfer Add PBS buffer with transfer pipette, empty wells with “flip and bang”  method. pipette, empty wells with “flip and bang”  method.

• Add the substrate:Add the substrate: Place 100 uL of the substrate Place 100 uL of the substrate in all 12 wells.in all 12 wells.

• IncubateIncubate at 37 at 37oo (oven on side bench) for 5 (oven on side bench) for 5 minutes.minutes.

• Evaluate the plate for color change Evaluate the plate for color change