B CELLS

Antibodies make up

-approximately 20% of normal plasma proteins. -Human serum usually has a broad gamma peak (gamma globulin) because normal individuals have a very diverse antibody profile (black line).

Haptens

-are small antigenic molecules that are NOT immunogenic by themselves. -frequently covalently conjugated to large carrier proteins. -The hapten-carrier complex is immunogenic and an anti-hapten antibody is generated. -Free hapten (non-conjugated) can bind to ab.

IgG amounts

-constitutes 75% of the total blood Ig in adults, and it is the major ab isotype found in the interstitial fluids. - 25% of the ab pool exchanges each day bw intravascular and extravascular sites. -so tissues to be constantly "bathed" in IgG containing plasma. - carried to tissues during inflam

Specificity

-describes the degree to which the ab differentiates antigens and can bind to one antigen and not another. -ab binds tightly to only one antigen leading to a high degree of specificity. -This is essential in targeting the destructive power of an immune response toward a foreign entity

IgG structure

-exists only as a monomer formed by 2 light chains and 2 gamma chains. -4 subclasses of IgG in humans: IgG1, IgG2, IgG3, and IgG4 encoded by g1, g2, g3, and g4 gene segments - made later during primary immune responses, and it is the major ab generated in most secondary immune responses. -IgG is usually higher affinity than IgM

Hx of B cell and ab

-idthe presence of a neutralizing substance in the blood that could counter infection. -passive immunization (transfer of antibodies from one individual to the next) has been used in clinical practice

Agglutination

-involves the clumping of particulate antigens such as bacteria or fungi. -Because abs have at least 2 antigen-binding regions, one ab can bind 2 epitopes on diff antigens and hold them together as a larger clump inhibiting its movement. - Abs can make the structure larger and increase the likelihood of increased phagocytosis by macrophages and neutrophils.

Neutralization

is a process by which an ab defects a cell or the host from an antigen or a pathogen. -Abs bind pathogens or toxins and sterically hinder their binding to cell surface receptors. -ex ab (IgA) often play a role in preventing attachment and colonization of mucosal sites - They are induced thru several vaccines (tetanus) to prevent the effects of microbial released toxins

paratope

is created by the light and heavy chain binding to the 3D conformation of the epitope

IgA in blood ad location

is present in blood and is the predominant antibody in seromucous secretions. -Monomeric and dimeric serum IgA constitutes 15-20% of blood immunoglobulin. -Secretory IgA (sIgA) is made at higher levels than all other Igs, and it is actively excreted through cells of the mucosal epithelium into secretions—saliva, tears, colostrum, and the bronchial, genitourinary, and intestinal tracts.

Papain digestion of IgG

leaves the molecule at the hinge region resulting in three fragments, two Fab fragments (antigen binding) and the Fc fragment (crystallizing / constant region; gives the antibody its effector function). -The Fc fragment contains the site(s) that allow the ab to bind to Fc receptors (FcR) on cells and to bind complement.

RNA Splicing

Roles: · Joining RNA exons and removal of introns · Accounts for IgM and IgD expression in same B cell (alternative RNA splicing) · Accounts for secreted vs. membrane-bound Ig forms (alternative RNA splicing/poly-A) Characteristics: · Allows expression of alternative pathways · Precise breakage/rejoining · Results predictable

DNA Changes

Roles: · VDJ gene rearrangement (antigen independent, 1 o lymphoid organ) · Isotype switching (antigen dependent, 2o lymphoid organ) · Somatic hypermutation in B cells (antigen dependent, 2o lymphoid organ) Characteristics: · Generally permanent · Often imprecise breakage/rejoining · Results in variability which is risky due to possible lack of function or autoimmunity

Chart IgE

normal sreum %: .004 half life: ~2-3 days (serum) months (mast cell) Strucutre: monomer Site: tissue mast cell • Mast cell degranulation • Parasites and allergy

Chart IgD

normal sreum %: .2 half life: ~2-3 days Strucutre: monomer Site: Naive B cells • Naïve B cell antigen receptor • Role unknown

Mast cell degranulation

can be caused by antigen cross-linking IgE already bound to high affinity FceRI receptors on mast cells and basophils

Features of antigens affecting immunogenicity: Adjuvants

can promote a robust immune response through nonspecific stimulation

affinity maturation.

causes The average affinity of ab molecules in a pop is substantially higher in secondary immune response than in primary responses.

Pepsin digestion of IgG

cleaves the molecule below the hinge region generating one major fragment, F(ab') that contains both antigen-binding regions.

Immunoglobulins consist of two major portions:

constant region: CH, Clambda, Ck variable region: formed by multiple gene segments coming together

Chart IgA

normal sreum %: 15 half life: ~6 days Strucutre: monomer and dimer (+J chain +/-S chain Site: Secretions (dimeric), Blood-mono & dimeric, Tissues - monomeric • Neutralization • Agglutination • Passive protection for infant via breast milk

Chart IgG

normal sreum %: 75 half life: ~21 days Strucutre: monomer Site: blood, tissues, fetus • Secondary immune response • Opsonin • Complement fixation + • ADCC by NK cells, M, PMNs • Neutralization • Agglutination +/- • Crosses the placenta

Chart: IgM

normal sreum %:~10 half life: ~10 days Strucutre: monomer (BCR) and pentamer (+ J chain) Site: Blood • Primary immune response • Naïve B cell antigen receptor • Complement activation ++++ † • Agglutination ++++ • Neutralization

Allelic exclusion

once a successful heavy chain (mu protein) is produced further rearrangement of the heavy chain is inhibited on the other chromosome · genes on the other chromosome will only rearrange if a non-productive rearrangement took place on the first chromosome · similar events occur to prevent expression of more than one light chain Allelic exclusion ensures: 1 receptor: 1 cell: 1 reactivity.

clonal selection theory

one clone of B cell produces only one antibody -further studies found the moleculare structure of Ab using multiple myeloma a cancer of plasma cell= 1 Ab

Framework regions

provide structural support for the HVs

Features of antigens affecting immunogenicity: Mode of administration

dose, route, and timing of administration are important in optimizing immunogenicity

hevay chain gene

encoded by a single gene locus on chromosome 14, and it determines the antibody isotype or class—IgM, IgG, IgE, IgA, or IgD. -The isotype produced is determined by which heavy chain constant gene segment is closest to the rearranged VDJ region on the gene. -The heavy chain gene segments for each antibody are identified by a Greek letter. m (mu) encodes IgM (heavy chain constant region) g (gamma) encodes IgG (heavy chain constant region) a (alpha) encodes IgA (heavy chain constant region) e (epsilon) encodes IgE (heavy chain constant region) d (delta) encodes IgD (heavy chain constant region)

Features of antigens affecting immunogenicity: Foreignness

generally, molecules must be recognized as non-self to generate an immune response, the more foreign the more immunogenic

Immunoglobulin molecules are composed of the following gene segments:

heavy chain: VH, DH, JH, CH (heavy chain locus) light chain: Vlambda, J lambda Clambda (lambda locus) -or- Vk, Jk, Ck (kappa locus)

Antibody-dependent cellular cytotoxicity (ADCC)

involves an innate effector cell which selectively kills a target cell coated with target antigen-specific Ig. -classic ADCC is mediated by NK cells, neutrophils, macrophages, eosinophils, and basophils can also undergo ADCC. -These cells express Fc receptors that allow them to bind to the Fc region. -The mech of killing/damage varies by the effector cell in question - multiple ab bind to bacterium and multiple Fc receptors

the result of gene rearrangement

10^9-10^11 different Igs are generated 10^7-10^10 different TCRs are generated The DNA in a mature B cell is unique from that in every other cell (even other B cells unless it arose from the same original clone). The genes encoding Igs and TCRs are DIFFERENT, and they are on DIFFERENT chromosomes.

basic structure of an antibody is comprised of

- 4 polypeptide chains, two identical heavy chains and two identical light chains. - this basic unit is termed a monomer. -the heavy chain is approximately twice the size of the light chain. -The heavy chain has 4 or 5 immunoglobulin domains -The chains are held together by intermolecular covalent disulfide bonds and non-covalent bonds in a bilateral symmetrical arrangement.

effector activites if IgG

- Activates complement via the classical pathway but requires 2 molecules so higher[ ] is needed for function - Opsonization (promotes phagocytosis) - Neutralizes - IgG crosses the placenta to provide passive protection to the newborn -Catabolism of IgG - one half-life every ~21-days -Passive protection in the newborn prevents many infections during the first months of life

IgG and affinity

- IgG is generally of higher affinity, a lower affinity IgM molecule can often bind tightly to a multivalent antigen by high avidity interactions. -the high affinity nature of an IgG interaction, the tissue distribution of IgG, and its extensive effector functions constitute the major advantages of a secondary immune response over a primary immune response.

IgE effector activities

- Mast cell and basophil degranulation - Important in defense against helminthic parasites (worms) - Associated with allergies and asthma

antigenic epitopes of B cells are created by

- a tertiary structure formed by a combination of the light and heavy chain. -Together they form the paratope--pocket or groove--that binds the 3-D structure of the epitope. -This interaction is exquisitely specific (lock and key fit).

immunogenic

- an immunogen binds to the antigen receptor and induces an immune response

light chain gene

- are encoded by genes found in the nuclear genome. -The light chain can be generated by 1 of 2 different loci--k (kappa) chain on chromosome 2 OR (lambda) chain on chromosome 22. -No functional differences have been identified between the k or lambda chains. -Each B cell expresses either k or lambda but never both. -This is called allelic exclusion.

B cells undergo development in the

- bone marrow, a primary lymphoid organ -.where they form their antigen receptor, the BCR, and gain their antigen specificity. -Naïve B cells emerge from the bone marrow and then travel through the body to one of many B cell secondary lymphoid organs. -(spleen/lymph node/MALT/bone marrow) where they wait for stimulation by antigen -Membrane-bound immunoglobulin (Ig) on the B cell binds epitopes on the antigen in a specific manner and signals the cell to be activated -Naïve B cells recognize antigen and other signals prompting them to proliferate and differentiate into short lived antibody-secreting plasma cells and long-lived memory cells. -"All" daughter cells and antibody created have the same antigenic specificity.

Junctional diversity

- changes in nucleotide sequences introduced at the junctions of V, (D), and J gene segments that DO NOT come from template DNA (de novo DNA sequences) a. Exonucleases remove nucleotides from V, (D), J gene segments b. An enzyme called terminal deoxyribonucleotidyl transferase (TdT) takes nucleotides that are not part of germ-line genes and adds them randomly to the sites of V(D)J recombination. These added nucleotides are called N-nucleotides.

Features of antigens affecting immunogenicity: Chemical and structural complexity

- epitope diversity contributes to the degree of immunogenicity: the more varied the epitope composition of an antigen, the more likely different (individual) immune responses will be induced.

Ig structure

- heavy chain is of the alpha class, and there are 2 subclasses—IgA1 and IgA2. -Dimeric (sometimes trimeric) are formed by monomers linked covalently by a J chain (like IgM). -Each sIgA (secretory IgA) also contains a secretory component. -The secretory component facilitates transport of the sIgA across the mucosal epithelium into the lumen and protects the sIgA from proteolysis. -sIgA is responsible for immunity at mucosal surfaces. - Consists of2 monomeric units plus a J-chain and a secretory component - Secretory component is synthesized by epithelial cells and makes it more resistant to proteolytic cleavage by enzymes commonly found in bodily fluids - sIgA protects mucosal surfaces by binding pathogens and interfering with their adherence and colonization (neutralization)

IgE

- is a monomer that comprises <0.004% of total blood Ig. - very short halflife in the blood (~6 hours), but it can remain bound to high affinity Fce receptors on mast cells for weeks to months. -IgE bound to its high affinity receptor is the only instance where immunoglobulins remain bound to their Fc receptors for long periods of time in the absence of antigen

Humoral immunity

- is mediated by antibodies and is the arm of the adaptive immune system that functions to neutralize and eliminate extracellular microbes and toxins. - B lymphocytes use membrane-bound antibodies to recognize a wide variety of antigens, including proteins, polysaccharides, lipids, and small chem. -These antigens may be expressed 1) on microbial surfaces (capsular components of bacteria or envelopes from viruses), 2) inside the microorganism (capsid or transcriptase from a virus), or 3) soluble microbial mediators (secreted toxins)

individuals with multiple myeloma (or Waldenstrom's macroglobulinemia) usually express

- predominantly a single antibody in their blood, and this shows up as a single tight peak (red line). - also have Bence Jones protein in their urine. -Bence Jones proteins are dimers of immunoglobulin k (kappa) or (lambda) light chains. -The use of antibodies from these individuals was critical to our understanding of Ig/ab protein structure and binding

isotype switching

- process:A single B cell (and heavy chain locus) has the capacity to make each of the isotypes -The heavy chain determines the antibody's effector activities and its tissue distribution. -allows the antibody to change its effector characteristics and distribution

secreted antibodies enter the circulation and mucosal fluids and bind to

- the antigens, leading to their neutralization and elimination. -The antigen receptors on B cells and the antibodies that are secreted usually recognize antigens in the native conformation with no requirement for antigen processing or specialized display. -The memory cells help the body to mount a more robust "secondary immune" response during subsequent antigen exposures.

Paratope features

- the region of the antibody that contacts the antigenic epitope · Composed of hypervariable regions (HV)/complementarity-determining regions (CDRs) (bind epitote) · HVs are short regions of ~10 amino with a highly variable amino acid sequence · Virtually every ab molecule contains a unique amino acid sequence in the HV region · Both the light and the heavy chain contain 3 HV regions

affinity maturation (result)

-- the binding affinity of ab generally increase with prolonged or repeated exposure to that antigen (as in a secondary immune response) promotes clearance of the pathogen facilitates the use of "smaller" abs (e.g., IgM -> IgG/IgA/IgE) -somatic hypermutation + antigenic selection -> affinity maturation -unlikely that the specificity of the ab (what the ab binds) will change due to somatic mutations -After hypermutation, the Ig can have either a higher or a lower affinity for its antigen. -Those with greater affinity for antigen are preferentially selected, while those that are lower cannot compete for antigen, and they die. - allows the affinity of abs to increase during the immune response. This process is called affinity maturation..

IgM

-10% of blood Ig. -Monomeric BCR on naïve B cells - first secreted ab made following activation of a naïve B cell; found early during a primary immune response - A single membrane-bound molecule of IgM is able to bind C1q allowing this antibody to function at very low titers/concentrations early in the response. -The BCR on naïve B cells surfaces is made up in part by monomeric IgM molecules. Effector activities - Most efficient at complement activation via the classical pathway - Neutralization - Agglutination Low affinity antibody with overall high avidity due to pentameric structure -, a lower affinity antibody that relies on avidity for binding

Why do we swithc isotypes

-As the B-cell response to infection gets underway, isotype switching diversifies ab function by changing the heavy-chain constant region. -The diff bw the isotypes serve several purposes. -By varying the # of antigen-binding sites and the flexibility of their movement, isotypic diff alter the strength with which ab bind to pathogens and antigens.

IgD

-D is a monomer. -It is synthesized to be a membrane-bound receptor, a BCR, on naïve B cells. -No secreted form of this antibody. -Free IgD is found in the blood at low levels after it "falls off" a B cell. -The role/requirement of IgD in biological function is unknown.

mmunoglobulins fall into five major isotypes or classes

-IgG, IgM, IgA, IgE and IgD. -These are distinguished based on structural differences in the constant part of the heavy chain. -These differences confer unique bio and chem properties that allow them to interact with a diff subset of immune-system proteins.

Ig M in the blood

-In the blood, it has a pentameric structure comprised of 5 IgM monomers made up of 2 light chains and 2 mu chains each. -The ab are then linked by a single J chain. -Blood IgM has 10 potential binding sites, but steric hindrance usually limits binding to 5 sites. -IgM is a pentamer,=robust activator of the classical cascade.

Details of the mech of gene rearrange

-The lymphocyte specific recombinases recognize specific DNA sequences (recombination signal sequences) located between the V and J segments and flanking both sides of the D regions. -These recognition sequences are highly conserved stretches of 7 (heptamer) and 9 (nonamer) nucleotides separated by non-conserved spacers. - the V(D)J recombinase brought the V and J exons into apposition, forming a "hairpin" loop of intervening DNA that is then randomly cleaved, TdT adds random nucleotides, and DNA double stranded break enzymes ligate the ends of the gene segments leading to the excision of the intervening DNA. -This results in the permanent loss of the excised DNA. -All clonally derived daughter cells will have the same change in their DNA and the same antigen specificity.

Antibody distribution (blood and tissues)

-ab are found in the blood plasma (the unclotted liquid component of blood), perfused tissues, and most secretions. -Ab isotypes are distributed selectively to each of these sites in the human body providing different types of immunoprotection as needed. - IgM and IgG are found in the blood while IgG constantly "bathes" the tissues as it exchanges between intravascular and extravascular sites. -sIgA is found in secretions and provides a barrier to microbial adhesion and colonization at mucosal sites. -IgE is found in mucosal tissues and connective tissues providing defense at those sites.

antigenic

-an antigen binds to an antigen recepetor, but it doesnt induce an immune response nessarily

binding of antigen to antibody takes place by the formation of

-multiple non-covalent bonds bw the antigen and AAs of the binding site. -Although the attractive forces involved in these bonds are individually weak by comparison with covalent bonds, the multiplicity of the bonds leads to the considerable binding energy. -The attractive forces binding antigen to ab include: electrostatic forces, hydrogen bonds, Van der Waals forces (flux e cloud), and hydrophobic forces

N-terminal domain

-of each heavy and light chain displays more variation in AA sequence than the other domains. -the N-terminal domain of each polypeptide chain is designated as the variable region and the remaining domains are designated as constant regions. - The variable region includes 2 distinct antigen binding sites formed by a combo of the light and heavy chains. -The variable region is made up of domains designated VH for the heavy chain and VL for the light chain • The sequence variation in VH and VL confers the antigen specificity of the antibody.

Membrane-bound immunoglobulins

-play a major role in B cell development and function. - membrane-bound Ig or the BCR is very important in signaling and activation of a B cell.

selective theory

-specialized cells (B cells) had receptors that bound a given pathogen this interaction induces the cell exhibiting the receptor to multiply and produce more copies of the same receptor

avidity

-strength with which a multivalent antibody binds a multivalent antigen -avidity is dependent on the underlying affinity of the interactions, it is a distinctive, stronger bond -When a multivalent antigen combines with more than one of an ab's combining sites, the binding energy between the 2 is considerably greater than the sum of individual sites since all the antigenantibody bonds must be broken simultaneously before the antigen and antibody dissociate - Ig has high avidity by high affinity -the total strength of the interaction bw the ab and the antigen, greater than the sum of all the affinities because all binding sites must dissociate at the same time for the antibody to release the antigen

The stages of B cell development

1) Stem cell commited to B cell lineage 2) Pro B cell ( heavy chain rearranges) 3) Pre B cells - heavy chan mu transcribed/ translated and in cytoplasm - light chain rearranges 4) Immature B cell- IgM expressed at cell surface, tolernace occurs here - Cell travel to 2nd lymphoid organs, matures and ecpresses IgD on surface 5) Mature NAive B cell- both IgM and IgD on cell surface 6) acivtated B cell - clonal prolif a) IgM secreting plasma cell and IgM synthesis b) B cell diff with T cell help I) IgG1 plasma and memory cells and IgG1 syntheisis II)IgG4 plasma and memory cells and IgG4 syntheisis III)IgE plasma and memory cells and IgE syntheisis IV)IgA plasma and memory cells and IgA syntheisis

several important differences between primary and secondary antibody responses:

1) The "lag-phase" or the time that separates the contact with the antigen and the appearance of specific abs. Abs appear much more rapidly during secondary or later responses. 2) The titer or amount of the ab response. Typically, secondary responses have several log factor higher responses. 3) The affinity of the ab for its epitope generally goes up with each antigen exposure. This process is called affinity maturation. 4) The predominant isotype generated in the BLOOD. During a primary response it is IgM early and IgG later. During subsequent exposures, it is predominantly IgG. Please note that sIgA is still the major isotype in SECRETIONS.

Creation of diversity

1. Multiple V, D, and J segments 2. Combinatorial diversity a. Different combinations of V, (D), and J gene segments b. Different heavy and light chains combining 3. Diversity gene segments (D) code in all three reading frames 4. Junctional diversity

Major consequences of gene rearrange

1. Only B cells express the Ig genes in a rearranged form 2. Occurs in the absence of antigen; therefore, naïve B cells (and T cells) exist prior to antigen stimulation which allows them to respond after antigen challenge 3. The process of Ig rearrangement is random and cannot be inherently biased towards recognition of microbes 4. Risks of DNA rearrangement · Failure to generate the appropriate antibody · A nonfunctional ("out of frame") receptor · Autoreactive receptors · Abnormalities in splicing (cancer)?

Things to remeber about isotype switching

1. The antigen specificity stays the SAME because the same VDJ region is "switched" from the original constant region to the new one 2. Somatic hypermutation occurs along with isotype switching 3. The light chain remains the same regardless of the antibody isotype

The BCR genes

1. The heavy chain is encoded by 1 gene 2. The light is encoded by 1 of 2 genes a. kappa light chain b. lambda light chain 3. There are only two copies of each of these genes (H, lambda and k)—one maternally and one paternally derived

Features of antigens affecting immunogenicity: Genetic constitution of the host

2 strains of the same species of animal or 2 individuals may respond differently to the same antigen. -Age, sex, and competency of the host may also influence the host response or lack of response to an antigen

what do B cell dervive from

B lymphocytes are derived from pluripotential stem cells in the bone marrow. -Development is independent of antigen but dependent on interactions with bone marrow stromal cells that provide a unique microenvironment for the successful early growth and differentiation of B cells. -Once committed to the B lineage, cells transit a series of developmental stages that are characterized based on the status of their Ig (heavy and light chain) genes. -If successful rearrangements at these loci are completed, it ends with an immature B cell that carries an antigen receptor in the form of a cell surface IgM that interacts with its environment.

. Gene rearrangement events create diversity

DNA rearrangements in the Ig gene loci occur in a precise order: 1. Heavy chain 2. Light chain a. kappa locus then b. lambda locus

How can the immune system react appropriately in each situation?

Different threats/pathogens require different responses for control and clearance. - the response to the same threat needs to be diff depending on where the threat/infection is and its stage. -recognizing a specific antigen allows the immune system to work in an appropriate, targeted manner against that threat. -Elimination of the threat results in the removal of the antigen that specifically drives the reaction and the down regulation of the adaptive immune response. -these cellular events promote a "memory" of the exposure. -This is done thru "residual immunity" including ab in serum and secretions and a pool of memory cells (B, TCTL, and TH).

Can a B cell make both IgG1 and IgE at the same time? Why/why not?

No bc need to cut out IgG1

A B cell is making IgA1. Can it undergo class switching to make IgG3?

No cut out alreaddy bc of isotype switiching

Heavy chain rearrangement

The heavy chain re-arranges first, with D-J joining following the deletion of intervening DNA. -After the D-J rearrangement, one of the many V genes is joined to the DJ complex, giving rise to a rearranged heavy chain gene. -Transcription and RNA processing leads to splicing of the C segment, giving rise to a functional mRNA for the mu heavy chain. -The phenotype of cell that only has mu(no light chain) is a pre-B cell.

Variable for B cells

The variable region of the light or heavy chain which includes either VDJ or VJ -or- The variable gene segment (V14, V22, or V75)

If two daughter cells isotype switch such that one is IgG2 and the other is IgA1 will they recognize the same antigen?

Yes bc VDJ is the same and this is used for gene recognition

Can a B cell make both IgM and IgD at the same time? Why/why not?

Yes bc alternative RNA splicing ( no Switch site)

isotype/class

a subdivision of Ig molecules based on structural and unique antigenic differences in the C regions of the H chains (Fc). -IgG subclasses, subdivisions of IgG, include IgG1, IgG2, IgG3, and IgG4. • gene segments within the heavy chain are exchanged by isotype switching

Complement fixation

abs are integral to the activation of the classical complement cascade. -The path is activated when C1q binds to sites on the Fc region of antigen-complexed IgG or IgM. -Complement activation can lead to pore formation in the target (a MAC complex), C3b binding and phagocytosis, and release of proinflammatory mediators (C5a, C3a).

Soluble immunoglobulins or antibodies

allow B cells to exert their effector function in the blood, extravascular spaces in the tissues, and multiple secretions (the humors)

How can a B cell make both a soluble AND a membrane form of IgM at the same time?

alternative RNA splicing

Antibodies

are protein molecules that recognize and bind to foreign substances (antigens) encountered in the environment. -comprise a class of proteins termed immunoglobulins, which are a heterogeneous group of glycoproteins that constitute approx 20% of total plasma proteins. - present in the circulation and on B cells

Light chain gene rearrangement

light chain locus and follows a similar sequence . -One V segment is joined to a J segment, forming the VJ complex (light chains do not contain D regions). -Splicing of the primary transcript joins the C gene to the VJ complex forming an RNA that is translated to produce a k or lambda protein. -The light chain is assembled with the previously synthesized mu chain in the ER to form the complete IgM molecule which is transported to the cell surface. -Naïve B cells make only membrane-bound forms of immunoglobulins.

The constant regions exhibit

little variation in AA sequence and have very similar secondary and tertiary structures. -A hinge region can be found in the constant region of many immunoglobulins (IgG & IgA). It is a flexible domain that joins the antigen binding arms to the Fc region. -The flexibility of the hinge region allows the antigen binding arms to adopt a wide range of angles, permitting binding to epitopes spaced variable distances apart. -This structure allows ab to carry out their dual tasks—binding on the one hand to a wide variety of antigens and binding a limited # of effector molecules and cells. • The constant regions are designated CL for the light chain and CH1, CH2, CH3, (CH4) for the heavy chain. • Some constant regions contain a hinge to facilitate binding to multivalent antigens.

A B cell is secreting IgM. What isotypes could it make after additional signals?

make all of them to IgA - cytokines drive change

B cell antigenic epitopes can be comprised of

many diff macromolecules including proteins, polysaccharides, lipids, nucleic acids, and many small chem. - B cell paratopes bind to epitopes of approx 4-5 AA or monosaccharide residues. -Bc the epitope is dependent on the 3-D structure, epitopes can be linear (continuous within the sequence) or conformational (due to protein folding). - This means that B cell ab have the potential to bind to soluble antigens, those that are an integral part of the membrane, or even degraded components. -allows ab to bind to just any potential threat.

Protein electrophoresis

method for separating proteins based on their size and electrical charge. -When serum is separated by electrophoresis, it forms a characteristic pattern of bands of different widths and intensities, reflecting the mix of proteins present. -This pattern is divided into 5 fractions, called albumin, alpha 1, alpha 2, beta, and gamma.

Cross-reactivity

refers to the ability of an individual ab combining site to react w/ more than one antigenic determinant or the ability of a pop of ab molecules to react with more than one antigen. -occur when Abs react with 2 molecules that share (identical or similar) epitopes but are otherwise dissimilar. -This can result in the failure of the immune system to distinguish "self" from "non-self" leading to host tissue damage or autoimmunity. -Some pathogens attempt to mimic host tissues to evade detection by the immune system this is molecular mimicry

lock-and-key theory

showing that the interactions bw ab and antigens depended more on their shape than on their chemcomposition

affinity

strength of a single antigen-ab bond is the ab -produced by the sum of the attractive and repulsive forces -.each monomeric ab has 2 antigen binding sites, abs have the potential to bind to more than one epitope on multimeric antigens, such as the repetitive components found on most pathogens. -the strength of a single site of antigen-antibody interaction, low affinity antibodies bind antigen weakly and tend to dissociate readily while high affinity antibodies tend to bind tightly to their epitope

Opsonization

the ab coats the surface of a pathogen to promote phagocytosis. -Antibodies (IgG) are a potent opsonin through the binding of their Fc region with Fc receptors on phagocytes. -Opsonins = immunoglobulins (Ig), complement (C3b), and acute phase proteins (CRP and MBP).

The solution for all the threats- gene rearrangement

the genes encoding the BCR undergo permanent changes in DNA sequence during B cell development to create millions of unique antigen receptors

somatic hypermutation (mechanism)

the nucleotides in the hypervariable regions of both the light and heavy chain undergo a very high rate of mutation -Once a B cell has been activated by antigen, further diversification can occur by a process called somatic hypermutation. -randomly changes single nucleotide bases in regions coding for the antigen binding domains.

Features of antigens affecting immunogenicity: Molecular size -

usually the larger the molecule, the better the immunogen. -Molecules <1,000 are only weakly immunogenic. -Certain small molecules (haptens) may be antigenic but not immunogenic unless coupled to a larger carrier protein. (They can be bound by antibody, but they are unable to activate adaptive immunity alone.)

Isotype switching from IgM to IgG/IgA/IgE

· IgM-producing B cells have not undergone isotype switching. · The VDJ region is adjacent to the first constant region, C mu, and the rest of the constant regions are 3' to Cmu · All the heavy chain constant regions are flanked by a switch site · The switch sites promote looping out of the DNA between the VDJ of the V region and next heavy chain C regions downstream and the intervening DNA is deleted. · Because DNA is lost, these are permanent changes in gene expression.

Isotype expression: IgM/IgD

· Naïve B cells (prior to antigen) express both membrane bound IgM and IgD and w/out isotype switching. · Activation of naïve B cells results in the production of the soluble pentameric form of IgM. · Membrane-bound IgM and IgD, and soluble IgM are all created by alternative RNA splicing, and/or alternative polyadenylation of transcripts generated from a single Ig heavy chain gene.

Antigen receptors (BCR or TCR)

· need to bind and distinguish between many, often closely related, chemical structures · usually recognizes one epitope on an antigen (1 lymphocyte: 1 receptor: 1 epitope) · all daughter cells produced from a common parental cell during clonal proliferation will also possess the SAME receptor. · a single B cell (or T cell) expresses ~100,000 identical copies of a single receptor

The molecular mechanisms of Ig gene rearrangement

· rearrangement of Ig genes occurs by excision (looping out) of the DNA between various gene segments · mediated by an enzyme complex, V(D)J recombinase composed of RAG-1 and RAG-2 (recombination-activating genes-1 and 2) · the RAG genes are specific for early stages of B (and T) cell development · mature proliferating lymphocytes and those producing ab generally do NOT change their functional specificity