General Introduction
• Specificity due to recognition of antigenic determinants or epitopes
• Epitopes = immunologically active regions that bind to:– Ag-specific membrane receptors on lymphocytes– Secreted antibodies
• Multiple epitopes can be found on single macromolecule
ANTIGENS = SUBSTANCES RECOGNIZED BY
• immunoglobulin receptor of B cells• T- cell receptor when complexed with MHC• Host determines whether B or T cell’s antigen-
binding receptor actually induces immune response
• B and T cell recognition are different
Immunogenicity and Antigenicity
• Immunogenicity = ability to induce humoral and/or cell- mediated immune response
• Antigenicity = ability to combine specifically with the final products of the above responses (i.e., antibodies and/or cell-surface receptors).
• Haptens, small molecules, are antigenic but incapable of inducing specific immune response
FACTORS INFLUENCING IMMUNOGENICITY
• Type of molecule: – Protein and polysaccharides immunogenic– Lipids and nucleic acids must be conjugated with
another molecule, i.e., like above
PROPERTIES DETERMINING IMMUNOGENICITY
Foreignness or non-self • Non-self antigens are eliminated by recognition
and response• lymphocytes that recognize self antigens are
eliminated by inactivation• Antigens that are more foreign are more
immunogenic– Exceptions:
• Collagen and cytochrome c – similar in all species• Corneal tissue and sperm - kept apart from immune system
Molecular Size• > 100,000 Da: excellent• < 5,000-10,000: poorChemical Composition and Heterogeneity• Homopolymers: poor• Protein structure: 1’, 2’, 3’, 4’ all contribute• Lipids serve as haptens Ab against lipids
useful – leukotrienes, steroids, vitamins
Susceptibility to Ag processing and presentation• Interaction of T-cells with processed Ag• Processing by: MΦs, neutrophils• Presentation via APCs (MΦs, DCs, B cells,
mast cells)• Larger molecules more easily phagocytosed
HOST CONTRIBUTES TO IMMUNOGENICITY
• Genes in MHC • Genes that encode B and T-cell receptors• Genes involved in immunoregulationEVIDENCE - Mice• Inbred strains of mice respond differently to Ag• F1 generation – intermediate• Backcross analysis mapped differences to MHC
subregion
HOST CONTRIBUTES TO IMMUNOGENICITY
EVIDENCE – Humans• HLA determines immune response to some
pathogens• African-Americans respond poorly to
interferon-α• HLA-DR4 associated with high responsiveness
to antigens specific to M tuberculosis but not to antigens shared with other mycobacteria (p = 0.0005)
HOST CONTRIBUTES TO IMMUNOGENICITY
Immunogen dosage• Dose response– Not enough to activate lymphocytes or tolerance– Too much tolerance
Example: capsular polysaccharide– O.5 mg: no IR– 0.0005 mg: Ab response
IMPORTANCE OF BOOSTERS: clonal proliferation
HOST CONTRIBUTES TO IMMUNOGENICITY
Immunogen route of administration• Routes: experimental Ag via intravenous (IV),
intradermal (ID), subcutaneous (SC), intramuscular (IM), intraperitoneal (IP)
• Influences which immune organs, tissues and cells – IV spleen– Subcutaneously local lymph nodes
HOST CONTRIBUTES TO IMMUNOGENICITY
Immunogen route of administration• Routes: vaccine Ag via intramuscular (IM),
oral, subcutaneous (SC),• Influences which immune organs, tissues and
cells – SC and IM local lymph nodes– Oral MALT
ROLE OF ADJUVANTS
ADJUVANTS = substances administered with Ag enhance immunogenicityUses: low immunogenicity OR only small amounts of Ag
availableMechanism of action: 1. Ligands for TLR on DCs and MΦ2. Prolong Ag persistence “depot effect”3. Enhance costimulatory molecules4. Increase local inflammation5. Stimulate nonspecific lymphocyte stimulation
ADJUVANT TYPES
HUMAN USE• Alum – aluminum potassium sulfate – APPROVED• MF59® – water in oil emulsion, Th1 and Th2 –
APPROVED in EU• AS04 - IN DEVELOPMENT: molecular mechanism based adjuvantsANIMAL USE ONLY• Freund’s incomplete adjuvantNOT USED EVEN IN ANIMALS• Freund’s complete adjuvant
EPITOPES
• immune cells do not interact with, or recognize, entire immunogen molecule
• Lymphocytes recognize discrete sites on immunogen molecule called epitopes, or antigenic determinants.
• Epitopes are the immunologically active regions of an immunogen that bind to antigen-specific membrane receptors on lymphocytes or to secreted antibodies
• Studies with small antigens reveal that B and T cells recognize different epitopes on same antigenic molecule
B-CELL EPITOPES
• Hydrophilic amino acids on protein surface– Protruding regions– Interior is hydrophobic; must be denatured to be
open to Ab• These recognize membrane bound or free Ab
• 15 - 22 amino acids on Ag contact Ab • 75–120 hydrogen bonds as well as by ionic and
hydrophobic interactions.
• Ag-Ab binding due to weak non-covalent interactions operating over short distances
• Precise complementary shapes increase non-covalent bonding
• Smaller ligands such as carbohydrates, small oligonucleotides, peptides, and haptens often bind within deep pocket of Ab
• Complex proteins contain multiple overlapping B-cell epitopes
• Only some are immunodominant• Determined with monoclonal antibodies
(MAbs)
• More potential antigenic sites than number recognized by immune system– Varies from species to species– Within species, individuals can • recognize different epitopes as immunogenic and• mount immune responses that are stronger
(immunodominant) against different epitopes
T CELL EPITOPES
• Antigen processing is required to generate peptides that interact specifically with MHC molecules
• Epitope is not conformational rather linear• Epitopes recognized by T cells are often
internal• given MHC molecule can selectively bind
variety of different peptides
• Antigen processed into antigenic peptides are presented in combination with MHC molecules
• Antigenic peptides recognized by T cells form trimolecular complexes with T-cell receptor and MHC molecule
HAPTENShaptens, small organic molecules that are antigenic but not immunogenic
Become immunogenic when linked to carrier molecule, e.g., large protein
drugs, peptide hormones, and steroid hormones
USE OF HAPTENS
• Configuration plays major role in determining whether it can react with a given antibody
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