Plasmid pHVDR22

1
Functional Assays
EBA (on plated cells)

• PvRII expressed on surface of plated mammalian
  cells with

Plasmid pHVDR22

Chitnis and Miller, 1994

• Plated cells are tested for binding with

- Human Duffy positive RBCs, and - Human Duffy
negative RBCs.
2
Functional Assays
EBA (on plated cells) Method

3
Functional Assays
EBA (on plated cells) Results
Comparative data for COS7 293Tcells Comparative data for COS7 293Tcells Comparative data for COS7 293Tcells
COS7 293T
Positive (fluorescent) 6 25
Binders per well 5.33 30.67
average of three experiments. average of three experiments. average of three experiments.

4
Functional Assays
Inhibition of Erythrocyte Binding
- 293T cells pHVDR22

• a-PvRII mouse serum

- Human RBCs

• 50 blocking at
• 125,000 dilution

• Blocking dependant
• on antibody dilution

5
Summary

• PvRII depends on Duffy antigen for RBC
  invasion,

• Blood-stage malaria vaccine candidate based on
  PvRII,

• Expression with baculovirus in insect cells,

• PvRII was found to be active in functional
  assays,

• PvRII was highly immunogenic,

• Anti-PvRII mouse sera (protein from insect
  cells), - recognises E.coli produced PvRII,
• - inhibits erythrocyte binding.

6
Acknowledgements
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