• Transfer of humoral immunity (pre-made Abs)
• Can occur naturally (breast milk) or artifically (antivenom or blood transfusions)
• Passive Immunity is NOT long lasting

• Igalpha/Igβ is needed for signaling after antigen binds BCR
• Analagous to CD3 for T cells
• Signaling motifis??

• Cross-linking is important for clustering of BCR and signaling molecules
• BCR on B cells cross linked by repetitive epitopes of antigens on the surface of a bacterial cells
• BCR on a mature, naive B cell is composed of surface IgM, which binds to antigen, and associated IgA/B chains which provide the signaling capacity

• Clustering of antigen receptors allows receptor-associated kinases (Blk, Fyn, or Lyn) to phosphorylate the ITAMs
• Syk binds to doubly phosphorylated ITAMs of IgB chain and is activated upon binding
• Syk can activate each other by transphosphorylation, thus initiating further signaling - induce changes in gene expression for B cell activation

• BCR crosslinking is neccessary but not sufficient to activate naive B cell
• Co-receptor composed of CD19, CD81, and CR2
• CR2 - complement receptor binding on pathogen
• CD19 - signaling component
• CD81 - unknown
• This simulatneous signaling strengthens the overall signal

• Binding of CR2 to C3d fragments on pathogen surface brings together the B-cell co-receptor with BCR
• Causes clustering on B-Cell surface
• Cytoplasmic tail of CD19 is then phosphorylated by tyrosine kinases associated with BCR
• Phosphorylated CD19 binds intracellular signaling molecules whose signals synergize with those generated by the BCR

• TI antigens; Thymus independent
• TI-1: example: LPS; acts by potentiating signal through TLRs - First signal here is Ag through BCR, then second signal is through TLRs
• TI-2: repetitive epitopes present at high density
• Only IgM produced, as cytokines (from CD4 T cells) are needed for isotype switching

• LPS is a TI-1 antigen that cativates LPS-specific B cell
• IgM antibodies specific for LPS
• LPS also activates B cells specific for other bacterial surface antigens in T-ind. manner
• IgM antibodies specific, for other components of the bacterial surface
• Bacterial DNA is a TI-1 antigen that can help activate B cells specific for bacterial surface growth
• IgM antibodies specific for bacterial surface antigen

B-cell activation by TI-2 antigen

• TI-2 binds B-cell co-receptor
• B-cell receptor signal for activation
• Activation

• B cells can encounter Ag in SLO and become activated (DC in lymph nodes) - induces a decrease in their mobility (trapped in T-cell zone of LN)
• They will then endocytose the Ag bound to their BCR
• This is processed and presented on MHC II
• Activated B cells will form conjugates with CD4 T cells specific for the same Ag

• Interaction of a B cells and T cells will cause CD40L to increase on the surface of B cell.
• This will interact with CD40 on the B cell that was also iduced upon activation
• Cytokines made by the activated, cognate T cell are also delivered to the B cell
• Drive B cell proliferation and differentiation into plasma cells

• After activation (Bc) in (Tc) area of LN, B:T cognjugates will form
• Move to medullary cords and undergos initial proliferation and secretion of IgM, which will enter lymph through efferent lymphatics - then activated B cells move to Bc area and the geminal center is induced

The primary and secondary foci for expanding antigen-activated B cells occurs at different sites in the LN

GERMINAL CENTER

• Somatic hypermutation and isotype switching happen in the geminal center.
• Site of massive proliferation
• End of proci germinal centers tend to be monoclonal or only have a few different clones of B cells within one germinal 

• Rapidly prolifeating Bc with no surface Ig. 1 division every 6hrs. These are the cells undergoing somatic hypermutation and class switching
• Give rise to centrocytes, which have surface Ig that are now mutated and class switched
• Cnetrocytes will die unless they interact with Ag and CD40L
• Move towards follicular dendritic cells (FDC) in germinal center for Ag interaction and then out toward mantle where T cells congragate at B:T cell border

• FDC have depots of antigen on their surface 
• Centrocyte will engage and endocytose their congnate Ag from FDC
• Complement activation covers virus particles with C3b - cleaved by factor I
• CR1/2 on FDC bind to intact virus particle and retain them at the cell surface

• Induce the expression of BCR (new spec.), -mutation random process
• High, Med., or Low affinity BCR made
• Needs to interact with cognate Ag at this point to survive
• No binding of Ag - die by apoptosis
• High affinity BCR - better chance of interacting with Ag on FDC - can differentiate into plasma cells
• Plasma cells can secrete Ab of diff. isotypes

• Centrocytes with identical high affinity Ig can differentiate into either plasma cells or memory Bc depending on the cytokines secreted by cognate Tch
• IL-10 - leads to PLASMA CELL (fight and terminate current infection)
• IL-4 - leads to memory B cells (future infections)

• Centrocytes with identical high affinity immunoglobulins can differentiate into either plasma cells or memory B cells
• IL-10 (plasma cells for fighting and terminating current infection)
• IL-4 (memory B cells for future infections)

• The cytokines produced and secreted by CD4 T cells influence the selection of a new heavy chain constant region for class switching
• CD40/CD40L interactions are also very important for class switch

Hyper IgM syndrome

• Can be caused by mutation in enzyme AID, as well as mutations in CD40/CD40L
• Cant undergo somatic hypermutation and cant isotype switch
• Produce mostly low affinity IgM
• Patients succumb to repeated bacterial infections
• Treated with serum from healthy donors

• Lymph node from patient with hyper-IgM syndrome (no germinal centers)
• Lymph node with germinal centers
• Hyper IgM syndrome can also be caused by mutation in or lack of CD40 or CD40L

Comparison of resting B Cells and plasma cells

• A plasma cell is terminally differentiated cell
• The Ag-specificity and isotype cannot be changed
• Resting Bc expresses antigen receptor in form of surface Ig - present to MHC II, can activate helper Tc
• Ig genes can also undego somatic hypermutation - progeny with altered Ig specificity
• Plasma - terminally differentiated Bc - synthesis/secretion of Ab

• FcR (receptor for constant fragment)
• FcεR (receptor for IgE)
• Transporting receptors and receptors for signaling/endocytosis
• Fluid-phase endocytosis of IgG from the blood by endothelial cells of BV
• Acidic pH of endocytic vesicle causes association of IgG with FcRn protecting from proteolysis
• Once reaching basolateral face of endothelial cell, the basic pH of extracellular fluid dissociates IgG 

• Binding of IgA to receptor on basolateral face of epithelial cell (ep/poly-Ig receptor/dimeric IgA/IgA secreting cell)
• Receptor mediated endocytosis of IgA (tight junction/basement membrane)
• Transport of IgA to apical face of epithelial cell (lumen/laminapropria)
• Receptor is cleaved, IgA is bound to mucus through the secretory piece (IgA dimer + secretory compon.)

• Use of FcRn for transport IgG across placenta (IgG)
• Use of pIgR to transport IgA to breast milk (dimeric IgA)

1. Repertoire Assembly
2. Negative Selection
3. Positive Selection 
4. Searching for Infection (Recirculation of Mature Bc)
5. Finding Infection (Activation/clonal expansion)
6. Attacking Infection (plasma cells/memory B cells)

• Mouse
• Stem cell (1,000-10,000)
• Mature B cell (>30,000,000)
• Mature T cell (>30,000,000)
• Lineage Selection: adoption of B cell characteristics
• Lineage Commitment: B cell signaling
• Ig Gene Rearrangement: VDJ rearrangement
• Homeostatis: stable numbers of B cell progenitors and mature Bc 

1. Induce specific transcription factors, drive gene critical to recombination, expand lymphoid progenittors - (HSC --> Lymphoid progenitor)
2. Heavy chain gene rearrangements, blocked light chain rearrangement, maintain B cell phenotype - (Lymphoid progenitor --> Late pro-B cell)

3. Stop heavy chain rearrangement, proliferate to make many cells, stop proliferating, rearrange light chain - (Late Pro-B cell --> Immature B cell)

4. Make sure that you: have a functional receptor, are not autoreactive, can survive when it leaves home - (Immature B cell --> Mature B cell)

• Every cell besides half of Immature B Cell and Mature B Cells are developed in the bone marrow
• Half of Immature B Cell and Mature B cell are developed in SLO

1. Pluripotent hematopoietic stem cell (CD34)
2. Common lymphoid progenitor (CD34 & CD10)
3. B-Cell precursor (CD34 & CD10 & CD127)
4. Pro-B-Cell (CD34 & CD10 & CD127 & CD19)

• Large pro B Cell
• u heavy chain is made
• Small pro B Cell
• u chain in ER
• Immature B Cell
• υ heavy chain
• gamma or kappa light chain
• IgM on surface

• Lymphoid Progenitor Cell - CAMs, VLA-4/VCAM-1
• Early Pro-B Cell - Kit/SFC
• Late Pro-B Cell - IL-7 receptor/IL-7
• Pro-B Cell - CAMs, IL-7/receptor
• Immature B Cell - IgM

Pro-B-Cell rearrangement of the heavy-chain locus is an inefficient process

• Productive Rearrangement
• D-J rearrangements on both chromosomes
• Signaled to survive and become pre-B Cells: 50% of cells
• Nonproductive Rearrangement
• V-DJ rearrangement on second chromosome
• Signaled to die by apoptosis: 50% cells

• Pro-BCR:
• VpreB 
• gamma5
• Both make surrogate light chain
• BCR:
• light chain assembled

• Allelic exclusion gives homogeneous BCR with high-avidity binding
• No allelic exclusion would give heterogeneous BCR with low-avidity binding

• Successive rearrangements are possible at the Ig light chain loci
• First VJ recombination
• Non-productive rearrangement
• Second VJ recombination
• Non-productive rearrangement
• Third VJ recombination

• Rearrangment 1st and 2nd chromosomes of kappa and gamma genes
• Produces IgM with kappa and IgM with gamma

1. First Checkpoint
• pre-BCR
• No pre-BCR = apoptosis
• Selects for functional heavy chains
2. Second Checkpoint
• BCR
• No BCR = apoptosis
• Selects for functional light chains

• Hematopoietic stem cell: locus is closed
• Early pro-B-cell specific transcription factors bind to Ig enhancers and promoters, opening up locus
• Late pro-B cel: D-J rearrangement is made
• Large pro-B cell: V-DJ rearrangement is made

• Chromosome 8  = MYC gene
• Chromosome 14 = Ig gene
• Translocation of portions can occur on one gene segement from both

1. Normal Chromosome 9 & Normal Chromosome 22 (BCR)
2. Chromosomes Break
3. Changed Chromosome 9 & 22 (philadelphia chromosome bcr-abl)

• Slide 20 BCD

• No reaction with self antigen
• No IgM binding
• Immature B cells moves to the blood and expresses IgD and IgM
• Reaction with self antigen 
• IgM binds
• Immature B cell is retained in bone marrow

• Central Tolerance (Bone Marrow)
• Receptor Editing
• Die by Apoptosis
• Become Anergic
• Peripheral Tolerance (Periphery)
• Becomes Anergic

• Self antigen ligates immature B cells Igm
• Immature B cell continues to rearrange light-chain genes

• Immature B cell makes a new light chain and thus an IgM with different specificity
• A.) If the new receptor is self-reactive, light-chain genes continue to rearrange
• B.) If the new receptor is not self-reactive the B cell leaves the bone marrow

A.) Successive new receptors are self-reactive. No further rearrangements are possible and the immature B cell undergoes apoptosis

• IgM of immature B cell binds soluble univalent self antigens
• B cell is signaled to make IgD and to become unresponsive to antigen
• Enters the peripheral circulation but does not survive for long

• Afferent lymphatic vessel and primary lymphoid follicle 
• HEV to B cell to Efferent lymphatic vesssel

• CCL21 attracts immature B cells to HEV
• CCL21 and CCL19 attacts B cells to LN
• CXCL13 attracts B cells into primary follicle
• Interactions with follicular DC and cytokines drives the maturation of immature B cells
• Mature B cells recirculate between lymph, blood, and secondary lymphoid tissues

• HEV - B cell/T cell - Germinal centers
• Germinal centers - plasma cells - efferent lymphatic vessel

• Slide 32

• Germline Theory: There exist enough genes in the genome to account for antibody diversity without involving novel mechanisms
• Instructional Theory: There exist only a few different antibody molecules but with distinct folding patterns depending upon the antigen contacted
• Somatic Theories: From a few genes encoding antibody H and L chains a wide diversity of Ag specificities could be obtained by gene recombination and/or somatic mutation
• 
  

• 1965 (Dreyer & Bennett): Ig H and L chains (2 genes) & Ig diversity resulted for V region and C region recombination
• 1976 (Tonegawa & Hozumi): identified gene recombination at the Ig locus 
• There are ~ 30,000 genes in the human body. However it can produce 3*10^11 different BCR/Ig

• Antigen-independent gene rearrangement
• BCR & TCR
• Antigen-dependent somatic hypermutation
• BCR only
• Antigen-dependent Ig heavy chain class switch
• BCR only
• Shuffles same Ag specificity to different effector molecules

• Light Chain - Chromosome 22 & 2
• Heavy Chain - Chromosome 14
• Slide 7 VDJ Recombination

• Alpha Chain
• LValphaX70-80
• JalphaX61
• Calpha
• Beta Chain
• LVbetaX52
• Dbeta 1 & 2
• Jbeta1X6 & Jbeta2X7 
• Cbeta1 & Cbeta2

• Of gene segments produces diversity in the antigen-binding sites of immunoglobulins
• Light Chain - Germline DNA --> Somatic Recombination --> VJ-join rearranged DNA
• Heavy Chain - Gerline DNA --> Somatic Recombination --> DJ joined DNA --> Somatic recombination --> VDJ joined DNA

• Recombination can only occur between BCR/TCR genes that have complementary RSSs
• This is how DNA is oriented for proper recombination ane religation
• Proteins associated with RSSs are required for recombination
• Read Slide 12

• Ubiquitous double-strand break-repair (DSBR) enzymes - (DNA-dependent protein kinase, Ku heterodimer, Artemis, and DNA ligase IV)
• Lymphoid-specific RAG-1/2 (RAG = recombinase activating gene)
• Lymphoid-specific terminal deoxynucleotidyl transferase (TdT)

Steps in VDJ recombination

Steps in DNA cleavage by RAG

1. Generation of junctional diversity
2. RAG complex cleaves the heptamer RSSs from the D and J segments to yeild DNA hairpins
3. RAG complex opens hairpins by nicking one strand of the DNA generating palindromic P-nucleotides
4. N-nucleotide additions by TdT
5. Pairing of strands
6. Unpaired nucleotides are removed by exonuclease
7. Gaps filled by DNA synth. and ligation (coding joint)

1. Germline DNA --> Recombination
2. Rearranged DNA --> Transcription splicing translation
3. Protein (TCR) 

1. Germline DNA --> Recombination
2. Rearranged DNA --> Translation splicing transcription
3. Protein (TCR)

• Expression of IgM - VDJ & Cu
• Expression of IgD - VDJ & Cdelta

• Junctional diversity from nucleotide addition increases the variability found within the CDR3 region of the TCR - this is the epitope/MHC binding region, thus generating more diverse TCR
• Two types of nucleotide additions: encoded or palindromic (P) additions and non-template encoded (N) or random additions

• Ig
• Less V gene pairs, less junctional diversity, less total diversity
• TCR
• More V gene pairs, More Junctional diversity, More total diversity

• In humans and mice, this is accomplished by gene recombination, along with random nucleotide additions/deletions at the gene joints
• In rabbits and chickens Ab diversity is obtained by gene conversion
• In cattle and sheep Ab diversity is largely acquired through somatic hypermutation

• Slide 25 VDJ
• Human Heavy-Chain
• Shark Heavy-Chain
• Light-Chain (Rays/Sharks)
• Chicken Heavy-Chain

1. Germline chicken Immunoglobulin genes (Chicken B progenitor RAG1/2)
2. Immature chicken B cells. All have rearranged the same Vh and Vgamma genes (All immature B cells in the bursa express the same receptor. Expression of slg induces proliferation
3. Sequences from V pseudogenes are introduced into rearranged V genes through gene conversion (Gene conversion creates variable receptor specificities - b cells without slg expression die)

• Somatice hypermutation targets the rearranged gene segments encoding the variable region
• Mutation frequency vs Transcription

• Day 7 Primary Response
• Day 14 Secondary Response
• Day 21 Tertiary Response
• As days increase Antibody affinity increases for CDR1, CDR2, CDR3 in the Heavy and Light Chain V regions

• Both have similar Ig gene segements Mouse 
• Mouse has Cgamma2b and Cgamma2a
• Human has different Ce and Calpha1 and Calpha2

• V-region assembly 
• Junctional diversity 
• Transcriptional activation
• Switch recombination
• Somatic hypermutation
• IgM, IgD expression on surface
• Membrane vs secreted form
• See SLIDE 31 VDJ

• X-linked severe combined immunodeficiency (SCID): first identified as a mutation in DNA-PKs; no B or T cells without Ag-specific receptor expression; "boy in bubble"
• Omenn Syndrome: mutations in RAG1 or RAG2 gene
• Irradiation-sensitive SCID: mutation in Artemis

• Mediated by neutrophils, NK cells, eosinophils, macrophages
• Use antibody to clear infection
• Fc receptor (FcR) binds to Fc region of Ab attached to target cells because of specific Ag-Ab recognition. This part is antigen specific, the action with the FcR is not
• Causes lytic enzymes and TNFalpha to release

1. Antibodies bind antigens on surface of target cells
2. Fc receptors on NK cells recognize bound antibody (FcgammaRIII or CD16)
3. Cross-linking of Fc receptors signals the NK cell to kill the target cell
4. Target cell dies by apoptosis

• Lytic enzymes (A)
• Perforin (B)
• TNF (C)
• Granzymes (D)
• Eosinophils (A & B) + Fc receptor
• NK Cell (B & C & D)
• Neutrophil (A)
• Macrophage (C & D) + Fc receptor

1. INF - early on for shutting down infected cells, helps initiate adaptive immune response (IFNa, IFNb, TNFa, IL-12)
2. NK cell mediated killing of infected cells - sense interferon and IL-12
3. T - cell mediated killing of infected cells

• Like T cells in that they kill target cells specifically but not through Ag specific receptors on the NK cells (no TCR or BCR)
• No memory formation
• Can kill/lyse target cells through same mechanisms as activated CD8 T cells - cytokines, inflammation response
• Innate immune cell - acts to condition environment and adaptive immunity by secretion of IL-12, IFNgamma
• NK-deficient people - severe varicella virus (CP) + CMV infection

1. Either NK cells recognize Ab coated cells through ligation of Ab Fc portion through FcR on NK cells - eg FcRgammaIII on NK cells will bind the Fc portion of IgG (ADCC)
2. Also can sense the amount of MHC I on a target cell
1. If MHC I is down reg. (occurs during certain viral infections [CMV, EBV], cancer) then the NK cell will not get an inhibition signal
2. If no inhibition signal NK cell will lyse target

• A target can give 2 different/opposite signals to NK cell
• One signal is activating, other is inhibitory
• Typically the inhibitory signal is mediated by engagement of MHC I on target
• If a negative signal is lacking, then the activating signal wins. Therefore ligation of the activation receptor = killing of target

• Lectin-like receptors and immunoglobulin-like receptors
• Killer immunoglobulin receptor (KIR) - mostly inhibitory
• C-type lectins - activating receptors - NKG2D
• Ligands on target cells are molecules associated with stress
• Altered self - low MHC I: activating signal

1. Normal Cell: MHC I  binding Inhibitory receptor, ligand binds activating receptor (+/-) NK cell
• No KILLING
2. Virus-infected cell: MHC I, ligand binds activating receptor - NK cell (+) 
• KILLING

• Activation
• MHC I on normal cells is recognized by inhibiotry receptors that inhibit signals from activating receptors
• NK cell does not kill the normal cell
• Inactivation
• Altered or absent MHC I cannot stimulate a (-) signal. The NK cell is triggered by signals from activating receptors
• Activated NK cell releases granule contents, inducing apoptosis in the target cell

1. Granzyme and perforin pathway: CTL and NK cells produce these molecule (QUICKER)
1. Perforin - pore forming
2. Granzyme - protease, induces DNA fragmentation both found in cytoplasmic granules in activated T cells (not naive) and resting NK cell
2. Fas - FasL pathway: Fasligand on activated Tcell 
1. Fas on Target cell (SLOWER)
2. 
   

• Conjugate Formation - receptor/ligand recognition and signaling - Granule
• Membrane Attack
• CTL Disassociation
• Target Cell Destruction by Apoptosis

• CTL - Target Cell --> Conjucate formation
• Granule - CTL-target cell conjugate --> CTL cytoplasmic rearrangement --> CTL granule exocytosis
• Dissociation
• CTL recycling

1. Ca2+ increases or crosses barrier
2. Granule
3. Perforin monomers
4. Perforin in target cell membrane
5. Polymerized perforin starts pore formation
6. Polymerized perforin - completed pore

• NK Cells, Macrophages, & Neutrophils
• FcgammaRIII (CD16) - gamma or squiggle
• FcgammaRIV - gamma or squiggle
• NK Cells
• NKG2C, D, E (CD94) binds DAP12
• Mast Cells & Basophils
• FceRI - alpha, beta, gamma

• Activating Receptors
• (KIR-2DS binds DAP-12 with ITAM)
• KIR3DS
• CD94 - NKG2C, E
• Inhibitory Receptors
• KIR-2DL - ITIM
• KIR-3DL
• CD94 - NKG2A, B

• IL-7 (receptor) - growth factor receptor
• stem cell, early pro, late pro, large pre
• RAG-1/2 - lymphoid specific recombinase
• early pro, late pro, large pre, small pre, immature, mature
• TdT - N-nucleotide addition
• early pro, late pro, large pre, small pre
• Gamma5 and VpreB - surrogate light chain comp.
• early pro, late pro, large pre, small pre
• Igalpha and Igbeta - signal transduction
• Early pro, late pro, large pre, small pre, immature b, mature b

1. Lymphoid progenitor cell - CAMs, VLA-4, VCAM-1
2. Early pro-B cell - Kit, SCF, bone marrow stromal cell
3. Late pro-B cell - IL-7, IL-7 receptor
4. Pre-B cell - CAMs, IL-7
5. Immature B cell - IgM

• Stem Cell - None
• Early Pro - None
• Late Pro - None
• Large Pre - u heavy chain is made
• Small Pre - u chain in ER
• Immature - u heavy chain, gamma or kappa light chain, IgM on surface

Pre B Cell (L-chain gene rearrangement)

Immature B Cell (Rearrangment Ceases)

• Slide 7 Complement

1. Central Tolerance: occurs in primary lymphoid organs - no reaction and immature B cell moves to the blood and expresses IgD and IgM
2. Negative Selection: B cells in Bone Marrow reacting with self Ag - retained in bone marrow

• Occurs if B cell reacts with multivalent self-Ag. Another light chain can be rearranged

• Clonal Selection: successive new receptors are self-reactive. No further rearrangements are possible and the immature B cell undergoes apoptosis

• Can develop after interaction with monovalent Ag
• These do not undergo receptor editing

1. IgM of immature B binds soluble univalent self g
2. B cell is signaled to make IgD and to become unresponsive to Ag
3. Enters the peripheral circulation but does not survive

• Negative selection occurs in the bone marrow and anergy can be induced in bone marrow; angergy early on in periphery
• There are no known mechanisms, like AIRE for T cells, whereby B cells become tolerant to Ag in other locations or inside cells, e.g. DNA

• The immature B cells are IgM+, but have little IgD
• Travel throught eh HEV into T cell area, then move to primary follicle, where final maturational events occur, through interaction with follicular dendritic cells (FDC) in primary follicles. FDC are not bone marrow-derived cell, are not related to DC or PDC
• Maturem naive B cells are IgM and IgD positive

Bone Marrow

Secondary Lymphoid Organs and Circulation

• Slide 14 Complement

1. Germline DNA - somatic recombination
2. Rearranged DNA - transcription
3. Primary RNA transcript - splicing
4. mRNA - translation
5. Polypeptide chain

• Part of innate immunity
• Not a PAMP
• Host Derived
• Binds to microbes
• Enhances phagocytosis, inflammation and therefore elimination of pathogen

• >10% of serum proteins ar C' proteins + >30 proteins, mostly made by liver
• Discovered because it helps in Ab mediated bacteria clearance (complementary)
• Normaly circulate in blood/serum in inactive forms
• In the presence of pathogen or Ab coated pathogens, C' becomes activated

• Multiple C' proteins can interact with each other to form different ways of killing pathogen directly or by induced phagocytosis
• Protease cascade, many C' proteins are zymogens, cleabing and activating each other
• The point is to have a bit bit of pathogen inducing a rapid and amplified response

1. Classical (activated by Ag-Ab complexes)
2. Alternative (activated by bacterial surfaces)
3. Lecting

• Activation releases major inflammatory molecules of the host

• Classical: C then #: C1, C2, etc
• When a cleavage reaction occurs, the products are designated by adding a lowercase letter
• C3 cleaved to C3a and C3b
• ALTERNATIVE pathway:
• Factor B, Factor D
• Factor B cleaved to Bb and Ba

• Found on APC to phagocytose pathogens
• This will also help to place pathogen proteins in MHC binding groove

• Mannose-binding lectin (MBL) and ficolins recognize and bind carbohydrates on pathogen surface
• MBL/ficolin, MASP-2
• C4
• C2

• C1q interacts with pathogen surface or with antibodies bound to surface
• C1q, C1r, C1s
• C4
• C2

• C3 undergoes spontaneous hydrolysis to C3(H2O) to initiate eventual deposition of C3 convertase on microbial surfaces
• Factor D & B
• Properdin - C3bbb
• C3 H2O

• C3a and C5a recruit phagocytic cells to the ssite of infection and promote inflammation
• Phagocytes with receptors for C3b engulf and destroy the pathogen
• Completion of the complement cascade leads to formation of a membrane-attack complex (MAC), which disrupts cell membrane and causes cell lysis

• MBL monomers form trimeric clusters of carbohydrate-recognition domains
• MBL binds with high avididty to mannose and fucose residues
• MASP-2, MASP-1
• Ficolins, similar to MBL in structure, have a different carbohydrate-binding domain
• Ficolins bind oligosaccharides containing acetylated sugars
• MASP-2, MASP-1

1. Associated with MBL or ficolin cleaves C4 to C4a and C4b, which binds to the microbial surface
2. C4b then binds C2, which is cleaved by MASP-2, to C2a and C2b, forming the C4b2a complex
3. C4b2a is an active C3 convertase, cleaving C3 to C3a and C3b, which binds to the microbial surface or to the convertase itself
4. One molecule of C4b2a can cleave up to 1000 molecules of C3 to C3b. Many C3b molecules bind to microbial surface

• MBL binds to mannose on pathogens, but not vertebrates
• MASP1
• MASP2
• Mannan polymer
• C3, C4, C2

• C3 --> Anaphylatoxin (C3a)
• C3 --> C3b
• C4b-C2a --> C3
• MASP1/2 --> Mannan polymer

• C1q - collagen region
• C1r
• C1s

• Pentameric IgM molecules bind to antigens on the bacterial surface and adopt the 'staple' form
• C1q binds to one bound IgM molecule
• Binding of C1q to Ig activates C1r, which cleaves and activates the serine protease C1s
• IgG molecules bind to antigens on the bacetrial surface
• C1q binds to at least two IgG molecules
• Binding of C1q to Ig activates C1r, which cleaves and activates the serine protease C1s

• C3 --> C3A or C3b
• C3b-Bb bound to properdin on LPS activated by Factor B and Factor D

• C3 - C3b
• Factor B (B) - Ba Bb
• Factor D (D) - D
• Properdin (P) - P

1. C3b deposited by classical or lectin pathway C3 convertase
2. C3b binds to factor B
3. Bound factor B is cleaved by plasma protease factor D into Ba and Bb
4. C3Bb complex is a C3 convertase, cleaving many C3 molecues to C3a and C3b

Aleternative Pathway

• Alternative pathway = "tickover"
• Catalyzed by spontaneous C3 cleabage
• Thus- does not depedn on pathogen-binding protein for its initiation
• C3 covertase = C3bBb
• Catalyzes additional cleavage of C3 (into C3a and C3b), leading to more C3bBb

• C5b --> MAC attack
• Anaphylatoxin & chemotaxic factor (C5a) activated by C5
• C5 activates C5b
• PMN binds C3b - C3b

• C5 convertase = C3b2Bb
• Leads to limited production of:
• C5b = MAC attack
• C5a = attracts neutrophils and monocytes, increase vascular permeability

• Deposit large quantities of C3b on the pathogen surface for removal via complement receptors

Classical, lectin and alternative pathways produce C3 and C5 convertases, just different ways

• C3 convertase:
• C3bBb (alternative)
• C4bC2a (lectin and classical)
• C5 convertase:
• C3b2Bb (alternative)
• C4b2a3b (lectin and classical)

• C5b binds to C6 and C7
• Forms partial pore
• C8 attaches next
• Followed by C9 forming complete pore

• Host cells (your cells) have complement regulatory proteins not found on pathogens, like DAF (decay accelerating factor) and MCP (membrane cofactor protein)
• DAF and MCP disrupt C3 convertase C3bBb on a human cell surface

• Sometimes elicited by antigens not associated with infectious agens, and this can cause disease

• Known generally as allergic reactions occur when a persons immune system reacts to inherently harmless 'environmental' antigens such as pollen, food, and drugs

• Substance that causes a reaction

• The tendency to be "hyperallergic" a predispostion to develop immune responses to common environmental antigens

• IgE
• Soluble antigen, Mast cell activation
• IgG
• Cell or matrix associated, complement FcR cells
• Cell surface receptor, antibody alters signaling
• IgG
• soluble antigen, complement phagocytes
• Th1/Th2/CTL
• Soluble (TH1/TH2), Marcrophage activation (1) IgE production, eosinophil activation (2)
• CTL - cell associated antigen - cytotoxicity

• Immediate - type allergic reactions
• Causes you to have symptoms very soon after Ag recognition
• Mediated by IgE
• Pollen 
• Food
• Venom
• Drugs

• Proteins, often with carbohydrate side chains
• Small & Highly soluble and resistant to digestion
• diffuse into the mucosa
• Occasionally proteolytic enzymes
• e.g. papain (from papayas) causes occupational allergy
• Usually delivered transepithelially at low dose
• favors Th2 responses and IgE production

• Sensitization
• Elicitation
• Slide 7

• Gastrointestinal tract
• Inc. fluid secretion, inc. peristalsis
• Expulsion of GI tract contents (diarrhea, vomit)
• Airways
• Decreased diameter, inc. mucus secretion
• Expulsion of airway contents (phlegm, cough)
• Blood vessels
• Inc. Blood flow, inc. permeability
• Edema, Inflammation, Inc. lymph flow and carriage of antigen to LN

• Systemic anaphylaxis: drugs, venoms, food, serum
• Acute urticaria (wheal-and-flare): animal hair, insect bites, allergy testing
• Seasonal rhinoconjunctivitis (hay fever): pollens, dust mites
• Asthma: danders, pollens, dust mites
• Food Allergy: Nuts, shellfish, dairy, soy, wheat

• Spotting test for allergens
• Sterile Needle - suspected allergen
• Postive test: area becomes red and swollen
• Number of suspected allergens are tested on the arm/back at the same time

• Most are palliative, not curative
• Antihistamines (e.g. hives), beta-antagonists (eg relax constricted muscle upon acute asthma attack), or general anti-inflammatories (eg topical corticorsteroids for eczema)
• Anaphylactic reactions treated with epinephrine (artificial adrenaline)

• Anti-IgE antibody (omalizumab)
• Attempt to restor tolerance to the allergen by injecting escalating doses of allergen (risk of inducing allergic response, but can be effective, may depend on inducing regulatory T cells)

• Results of four cross-sectional surveys of school children
• Drastic increase from 1990- present in asthma and eczema

• Atopy is increasing in Western industrialized countries
• Up to 40% show exaggerated tendency to mount IgE to common environmental Ags
• Hypothesis: decreased early exposure to certain infections or commensals increases atopy
• People who live on farms, have pets have fewer allergies

• Genetic - High (Atopic) - Low (Non-atopic)
• Environment (early exposure to ubiquitious microorganisms Helminth infection Hepatitis A virus Composition of gut commensal microbiota
• Less hygienic (non-atopic)
• Hygienic (atopic)

• Asthma more prevelant in developed countries over non-developed (>10%)

• IgG dependent response to cell associated Ags
• e.g. antibody-mediated destruction of RBC (hemolytic anemia) or platelets (thrombocytopenia) can be caused by some drugs (the antibiotics penicillin and cephalosporin)
• drug binds to cell surface and serves as target for anti-drug IgG, causes clearance of the cell by FcgR+ tissure macrophages (unclear why some ind. mount anti-drug IgG)

• IgG dependent response to soluble Ags
• Pathology is caused by the deposition of Ag:Ab aggregates (immune complexes)
• large aggregates are readily cleared from circulation
• However, small complexes (that form when Ag is in excess) tend to be deposited in blood vessel walls

• Inhaled allegens provoke IgG responses instead of IgE responses - caused by repeated high does exposure to hay dust or mold spores, leading to immune complexes within walls of alveoli, local inflammation, compromised lung function, and even permanent damage

• Immune complexes activate local leukocytes including mast cells (e.g. through FcgRIII), induce vasodilation, inflammatory infiltrate, complement activation

• systemic hypersensitivity reaction
• results form the injection of large quantities of Ag
• Freq. followed the administration of therapeutic horse antiserum (e.g. antivenin & and anti-pneumococcal Abs used in pre-antibiotic days)
• Anti-TNFa monoclonal Ab therapy is associated with serum sickness in some patients
• Serum sickness can cause tissue injury
• Symptoms - chills, fever, urticaria, arthritis, and sometimes glomerulonephritis

• Foreign Serum Injection - High to decreasing
• Antigen:Antibody Complexes (Fever, Vasculitis, Arthritis, Nephritis) Sharp high - sharp decrease
• Antibody against foreign serum proteins - none to gradually increasing 

• DTH (delayed type hypersensitivity)(as opposed to immediate hypersensitivity as you would have in Type I reaction)
• T cell dependent response
• TH1 to soluble Ags
• TH2 to soluble Ags - chronic asthma
• CTL to cell-associated Ags (poison ivy)

• Delayed-type hypersensitivity
• Proteins: Insect venom, mycobacterial proteins - local swelling, erythema, induration, cellular infiltrate dermititis
• Contact hypersensitivity
• Haptens: poison ivy, metal ions - local epidermal reactions, erythema, cellular infiltrate vesicles, intraepidermal absccesses
• Gluten-sensitive enteropathy (celiac disease)
• Gliadin - villous atrophy in small bowel, malabsorption

• A prototypic DTH response mediated by TH1
• Small amounts of Ags from M. tuberculosis are injected intra-dermally
• In people that have been exposed through infection or immunization a local T cell mediated inflammatory reaction evolves over 24-72hrs
• TH1 cells recognize peptide:MHC II complexes, release inflammatory cytokines, induce vasodilation and visible swelling

DTH response to injected Ag (e.g. TB test)

1. Ag is injected into subcutaneous tissue and processed by local antigen-presenting cells
2. A TH1 effector cell recognizes Ag and releases cytokines, which act on vascular endothelium
3. Recruitment of phagocytes and plasma to site of Ag injection causes visible lesion

• Similar to reaction to injected Ag, but caused soley by skin contact
• Ags include highly reactive small molecules that penetrate skin, particularly those that cause itching and scratching
• E.g. chemicals then react with self-proteins, and form hapten:protein complexes that may be presented on MHC molecules and recognized by TH1 or CD8 T cell as foreign Ags

• Sensitization Phase (Afferent Phase)
• Elicitation Phase (Challenge Phaes, Efferent Phase)

1. Skin (sensitization site) - langerhans cells & hapten
2. Afferent Lymph (Draining Lymph Node)
3. Effector CD8+ T cells and Regulatory CD4+ T cells
4. Efferent lymph
5. Skin (Challenge Site)

• Rash produced by contact with poison ivy is common example of allergic contact dermatitis
• CD8 T cell respnse induced by urushiol oil in the plant
• This oil contains lipid soluble chemicals that cross the cell membrane and attach to intracellular proteins, which form neo-antigens that are presented on MHC I
• Once sensitized, CD8 T cells kill host cells and secrete cytokines upon re-exposure

• Test panels. The individual packages are opened and applied to the back. Removed 48 hrs later

• Nickle Sulfate - 14.2%
• Neomycin sulfate - 13.1%
• Fragrance mix - 11.7%
• Balsam of Peru - 11.8%
• Colbalt dichloride - 9.0%
• Formaldehyde - 9.3%
• Quaternium-15 - 9.0%
• Thimerosal - 10.9%