INMUNOLOGIA MOLECULAR I.ppt [Sólo lectura]Microsoft PowerPoint - INMUNOLOGIA MOLECULAR I.ppt [Sólo...
Transcript of INMUNOLOGIA MOLECULAR I.ppt [Sólo lectura]Microsoft PowerPoint - INMUNOLOGIA MOLECULAR I.ppt [Sólo...
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INMUNOLOGIA MOLECULAR
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HUMORAL CELULAR
INMUNIDAD IgM MacrófagosNATURAL Células NK
(sin memoria) Neutrófilos
INMUNIDAD IgG Linfocitos CD4+ ADAPTATIVA (helper)(con memoria) Linfocitos CD8+ (CTL)
JM/03Medicina Molecular
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Pathways of differentiation of a pluripotenthematopoietic stem cell of the bone marrow
136380
differentiate in the thymus
differentiate in the bone marrow
Stemcells
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ANTIGENOS
Moléculas reconocidas por receptores en células B (B cellreceptor) o T (T cell receptor).
Hapteno : antígeno que para ser reconocido debe ser unido a un
carrier (hidratos de carbono)
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How many environmental antigens are we exposed to
during our lifetime?
The immune system responds to hundreds of thousands of foreign antigens introduced from the environment
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Examples of antigen, immunogen and epitopes
Antigen
Epitope A Epitope B
Immunogen
Epitope B
Antibody A Antibody B
Antibody B
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ANTICUERPOS
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RECEPTORES ANTIGENICOS
• B cell receptor (linfocitos B) = reconoce Ag nativo
• T cell receptor (linfocitos T) = reconoce Ag procesado
JM/00Medicina Molecular
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ANTIGENO CON MULTIPLES EPITOPES
SUERO CON MULTIPLES ANTICUERPOS
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APCB Ab
CD4
CD8
Ag
Ag Processing
HelpHelp
1. 2.
The Immune Response:
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LINFOCITO CD4 ACTIVADO
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Role of T-Helper cells in antibody formation.
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LINFOCITO B EN REPOSO
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LINFOBLASTOS B
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Lymph node
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Germinal center in lymph node
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PLASMOCITO
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PLASMOCITO APOPTOTICO
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Primary and secondary antibody responses.
136384
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¿Cómo se produce la gran diversidad de anticuerpos?
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Susumu TonegawaPremio Nobel 1987
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Producción de cadena κ
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GENES CODIFICANTES PARA INMUNOGLOBULINAS
• Cluster IGH: cadena H; crom.14;V,D,J,C.
• Cluster IGK: cadena K; crom.2; V,J,C.
• Cluster IGL(λ): cadena λ; crom. 22; V,J,C.
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ANTICUERPOS MONOCLONALES
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b. 1927(in Bahia Blanca, Argentina)d. 2002
b. 1946d. 1995
b. 1911d. 1994
MRC Laboratory of Molecular BiologyCambridge, United Kingdom
Basel Institute for ImmunologyBasel, Switzerland
Basel Institute for ImmunologyBasel, Switzerland
Argentina and United KingdomFederal Republic of GermanyDenmark
1/3 of the prize1/3 of the prize1/3 of the prize
César MilsteinGeorges J.F. KöhlerNiels K. Jerne
Find a Laureate:
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• antibodies produced by differentiated B-cells
• recombination results in B-cell lineages producing distinct antibodies
• serum contains antibodies representing many lineages
• difficult to isolate B-cells of defined specificity
• cannot grow B-cells in culture
Kohler and Milstein (1975)
• fused B-cells with myelomacells stable hybridoma
• hybridoma secretes monoclonal antibody
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• immunized mouse making desired antibodies
• boost 3-4 days before fusion• remove spleen and harvest
cells• mix with myeloma cells in
presence of fusogenic agent
Balb/c Mouse
MOPC 21 (tumor)
P3K (cell line)
P3-X63Ag8 (HGPRT-)
X63-Ag8.653 (IgG-)
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HGPRT = hypoxanthine-guanine phosphoribosyl transferaseDHFR = dihydrofolate reductase
HAT Medium:• Hypoxanthine
(purine salvage)• Aminopterin
(DHFR inhibitor)• Thymidine
(pyrimidine salvage)
Nucleotide Metabolism:
• salvage and de novo pathways
• HGPRT essential for purine salvage
• DHFR essential for de novo synthesis
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Selection in HAT Medium
Human mAbs produced by trans-forming lymphocytes with EBV
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Cloning Methods•soft agar• limiting dilution
Screening/Cloning
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• produce mAbs in vitro or with ascites
• harvest culture media (supernatant)– in vitro material is less
concentrated and contains bovine serum
• ascites are tumors grown in peritoneal cavity– ascites fluid and sera
contain high [mAb]– minor contamination
with mouse IgG
Production
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HAMA ("human anti-mouse antibodies"). HAMA ("human anti-mouse antibodies").
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Mouse monoclonal antibodies have been genetically engineered to replace all of the antibody molecule with human counterparts except the hypervariableregions directly involved with antigen binding.Humanized monoclonal antibodies are
currently be tested in human clinical trials.
Humanized Monoclonal Antibodies
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RECEPTORES ANTIGENICOS
• T cell receptor (linfocitos T) = reconoce Agprocesado
• B cell receptor (linfocitos B) = reconoce Agnativo
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MECANISMO DE ACCION DE LOS ANTICUERPOS
Via ComplementoADCC (antibody-dependent-
cellular-cytotoxicity
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• COMPLEMENTO
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MAC (complejo ataque complemento)