IMSE 311: Antibody & B-Lymphocytes

B-cell Progenitor (Pro-B Cell)

• Earliest development stage

• Growth Factor: (secreted by bone marrow stromal cells)

  • IL-7

• Transcription Factors:

  • E2A

  • Early B-cell factor (EBF)

  • Interferon regulatory factor 8 (IFR8)

  • Paired box protein 5 (PAX5)

• Rearrangement of B-cell receptor (BCR) genes: cell surface version of an Ig/Ab molecule that code for heavy and light chains of Ab molecule

• Markers:

  • CD19

  • CD45R

  • CD43

  • TdT

  • RAG-1

  • RAG-2 enzyme

B-cell Precursor (Pre-B Cell)

• Pro-B cell enters Pre-B cell stage after BCR gene rearrangement

• Markers:

  • Pre-BCR: heavy chains + surrogate light chain

Pre-B Cell Receptor (Pre-BCR)

Formed when a heavy chain combines with a surrogate light chain and signal-transducing subunits (Ig-alpha & Ig-beta) on B-cell surface

Light Chain Gene Rearrangement

Begins when pre-BCRs appear on the B-cell surface

Disulfide bonds

Combines heavy and light chains (immunoglobulins “Ig”)

Immature B-cell

• Preparation for antigen-specific response

• Marker:

  • IgM BCR

  • CD21

  • CD40

  • Class II MHC

Central Tolerance

Elimination of B-cells that bear self-reactive receptors

• more than 90% of B-cells die in this manner

Mature B-cell / Naive B-cell

• In the spleen

  • Follicular B-cell: recirculate between blood and secondary lymphoid organs in search of specific Ag (Majority of immature B-cells become follicular)

  • Marginal Zone B-cell: reside mainly in spleen’s marginal zone and respond rapidly to blood-borne pathogens

• Markers:

  • Complete IgM and IgD BCRs

  • Class II MHC

Plasma Cells

• Fully differentiated lymphocyte for antibody production

• Minimal surface Ig

• Abundant cytoplasmic Ig

• Marker:

  • CD138

Immunoglobulin

Glycoproteins produced by plasma cells found on B-cell surface that serve as receptors that specifically recognize foreign antigens to initiate the immune response

• Neutralize toxic substances

• Facilitates phagocytosis

• Composed of 2 heavy chains and 2 light chains (H2L2) “tetrapeptide”

Valence

Number of antigen-binding sites of an antibody (connected to Fab fragments)

• most Ab are bivalent (2 binding sites)

Fab Fragments

Located at at the amino-terminal end and have antigen-binding capability

• Consists of

  • 1 light chain

  • 1/2 heavy chain

Fc Fragment

Located at the carboxyl-terminal end and are responsible for important biological activities

• Spontaneously crystallized at 4° C

• Consists of

  • 2 heavy chains halves

Papain

Proteolytic enzyme that cleaves the Ab molecule above disulfide bond, yielding 3 products

• 2 Fab

• Fc

Pepsin

Proteolytic enzyme that cleaves at the carboxyl-terminal side of interchain disulfide bonds, yielding 2 products

• F(ab)₂

• Fc

Antibodies Acc. to Sedimentation + Molecular Weight

S = Svedberg unit (how fast a molecule sediments in a centrifugal field; positive relation to MW)

(high S means faster sedimentation) M E D A G

• IgM – 19 S (900 kDa)

• IgE – 8 S (190 kDa)

• IgG – 7 S (180 kDa)

• IgA – 7 S (160 kDa)

• IgD – 7 S (150 kDa)

Antibodies Acc. to Abundance

G A M D E

1. IgG: 70–75%

2. IgA: 10–15%

3. IgM: 5–10%

4. IgD: 0.001%

5. IgE: 0.0005%

Antibodies Acc. to Formation

M D G A E

1. IgM

2. IgD

3. IgG

4. IgA

5. IgE

Hinge Region

Flexible part of the antibody located between CH1 and CH2 region

• Determines half-life of antibody

• Larger hinge region = more flexibility = shorter half-life

• Ig G-A-D: antibodies with a hinge region

• Ig M-E: antibodies with no hinge region

Light Chain

Where antigens bind

• MW: 25 kDa

• Types: (2)

  • kappa

  • lambda

• Regions: (2)

  • 1 constant: structural integrity (carboxyl-terminal end)

  • 1 variable: forms antigen-binding site, together with variable region of heavy chain (amino-terminal end)

Heavy Chain

Effector functions

• MW: 50 kDa

• Types: (5)

  • gamma

  • mu

  • alpha

  • delta

  • epsilon

• Regions (4)

  • 3 constant: determines antibody isotype (carboxyl-terminal end)

  • 1 variable: forms antigen-binding site, together with variable region of light chain (amino-terminal end)

IgG

• Monomer

• 75-80% (most abundant)

• 23-25 days (longest half-life)

• New born immunity (can cross placenta)

• Starts complement pathway

  • IgG1 - large hinge region

  • IgG2 - short hinge region, least effective in crossing placenta

  • IgG3 - largest hinge region

  • IgG4 - short hinge region

IgA

• Dimer

• 10-15%

• 6 days

• Patrol mucosal surfaces

• Protection for infant digestive tract

  • IgA1 - mainly in serum, monomer

  • IgA2 - mainly in mucus membranes, dimer with J chain

IgM

• Pentamer (in body fluids), Monomer (on B-cell surface)

• 5-10%

• 6-10 days

• First responder

• Starts complement pathway (most effective in classical complement pathway)

• Agglutination and Opsonization

IgD

• Monomer

• 0.001%

• 1-3 days

• Heat labile

• Second to appear

• B (memory) cell production

IgE

• Monomer

• 0.0005% (least abundant)

• 2-3 days (longest half-life)

• Most heat labile

• Found on BASOs and MASTs

• Triggering of acute inflammatory reaction (allergy)

• Destruction of large antigen such as helminths