Transcript 1.9mb ppt - UCLA.edu

T/B Collaboration
Associative recognition/carrier effect
Hapten cannot elicit an immune response without a carrier
Hapten must be physically linked to carrier to generate response
Secondary immunization must be with the same hapten-carrier conjugate
for a memory response to occur.
1º Immunization
DNP-BSA
DNP + BSA
DNP-BSA
DNP-BSA
DNP-BSA + BGG
2º Immunization
DNP-BSA
DNP + BSA
DNP + BSA
DNP-BGG
DNP-BGG
2º 
 -DNP PFC
(Plaqueforming cells)
+
+*
*Carrier effect can be circumvented by priming separately with
The unrelated carrier used in the 2o immunization
In vivo Injection of Hapten/Carriers
DNP-BGG
1)
OVA
7 days
2)
2)
3) Ag
Ag
DNP-BGG
OVA
DNP-OVA
DNP-KLH
DNP-KLH + OVA
4)
DNP Ab
++++
++++
+
+
7 days
measure
Ab resp
Hapten and carrier need to be physically
Linked to allow T/B interactions and
observe responses
Interpretation: Ag “bridges” T and B cells
T/B Collaboration in vitro
Cells added to upper or lower chamber
In the presence of APCs
U pper:
None
None
None
TKLH
TKLH
Low er
TKLH+B DNP-KLH
TKLH + B DNP-KLH
B DNP-KLH
B DNP-KLH
B DNP-KLH
Ag
none
DNP-KLH
None
None
DNP-KLH
T and B cells can interact at a distance in vitro
This is accomplished by soluble cytokines
Close proximity of cells eliminates need for Ag “bridge”
DN P Ab:
59
960
72
84
1011
MHC Restriction at the Ab production Level
Bone Marrow Chimeras
Lethally
irradiate
B
B
BM from B saves it
A
BM from A does not save
Make BM chimera, wait 3 months, remove splenocytes
Incubate in vitro with TNP-KLH and look for primary response
A
B
B
A
A AxB
(A+B)
B
(AxB)
B
TNP Ab produced
0
0
+++
+++
+++
T cell help for B cells
is restricted
MHC II on APCs must be
compatible
Bone Marrow Chimeras
BM from B
B mouse
B strain T cells
Restricted
To B haplotype
BM from A
A strain T cells
Restricted
To B haplotype
Help
provided
-KLH
CD4
-TNP B
-KLH
CD4
TNP-KLH
MHC
II
Mf
B mouse
MHC
MHC
II
II
Mf
X
-TNP B
MHC Restriction
MHC X
MHC X
a-A
a-A
T cell
T cell
MHC X
a-A
T cell
MHC
X
MHC
APC
RECOGNITION
Ag B
Ag A
Ag A
APC
NO RECOGNITION
Y
MHC
APC
NO RECOGNITION
A T CELL WILL ONLY RECOGNIZE A SPECIFIC PEPTIDE IN THE CONTEXT OF ITS OWN MHC MOLECULE
X
CTL will
Only kill
Infected
Cells of
The correct
(self)
haplotype
Specificity
Defined by
Both peptide
And MHC
MHC I
Variability
Dual-Reactive Hybridoma
Reacts w/ 2 different Cyt C molecules
Same MHC restriction (IEk)
Make point mutants in MHC II (APC)
Look for IL-2 production by T cell
AA of MHC II bind TCR in a
Different pattern if peptide different
Similar changes if mutate one AA in
TCR V or Vb
TCR/MHC-peptide interactions
Are fluid. Different conformations
May influence affinity of binding.
Alloreactivity
1 - 10% of T cells will react against an allogeneic stimulus
Explained by Cross-reactivity
1. Peptides bound by allo APC (but not self)
Form complexes self T cells not tolerized to, self T cells will react
Allo peptide strongly binds TCR that normally binds self MHC + different peptide
2. Direct binding by self T cells to distinct features on allo MHC
Independent of peptide (although peptide must be present in Ag cleft)
High concentration of foreign MHC creates a strong signal
ALLOREACTIVITY
Ag + self MHC
Binding
T CELL
Peptide-Dominant
Binding
MHC-Dominant
Binding
T CELL
Self
Class II
T CELL
Foreign
Class II
APC
APC
ALLOREACTIVITY COULD BE CAUSED BY CROSS-REACTIVITY W ITH PEPTIDE (MIDDLE PANEL)
OR BY CROSS-REACTIVITY W ITH FOREIGN MHC MOLECULE
APC