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Title: Development and characterisation of anti-DBLβ surface-labelling and cytoadhesion-inhibitory mouse monoclonal and polyclonal antibodies
Author: Alkurbi, Mohammad
ISNI:       0000 0004 5369 2020
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2016
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Plasmodium falciparum is responsible for most malaria-related morbidity and mortality, mostly affecting young children, non-immune adults and pregnant women. A characteristic feature of the pathogenesis of infection caused by P. falciparum is the cytoadherence of infected erythrocytes to the endothelial cells lining the microvessels of host organs. This phenomenon, termed ''sequestration'', mainly results from the adhesive interactions between P. falciparum erythrocyte membrane protein-1 (PfEMP1) proteins on the surface of infected erythrocytes and various host endothelial receptors such intercellular adhesion molecule 1 (ICAM-1), which is hypothesised to have a role in cerebral malaria. PfEMP1 molecules consist of several Duffy binding-like (DBL) and cysteine rich interdomain region (CIDR) domains that have different cytoadhesive functions. The second class of DBL domains, DBLβ, has been associated with adhesion to ICAM-1 receptors. In the present study, we selected four recombinant PfEMP1ICAM-1-DBLβ domains for mouse immunisations. Thirteen monoclonal (mAbs) and polyclonal antibodies (pAbs) were raised to three recombinant domains (DBL13, DBL31 and DBL41). All mouse mAbs and pAbs comprised IgM antibodies that recognised homologous and heterologous DBLβ domains. Most mAbs and pAbs labelled the surface of erythrocytes infected by P. falciparum isolates, with an IgM labelling capacity ranging from 10.1% to 67.6% of total IEs. Mouse antibodies showed similar patterns of reactivity with ICAM-1-binding and non-binding isolates, and reacted with a parasite isolate from a different genome (3D7). Surprisingly, we detected a remarkable reduction in IE population after incubation with mouse mAbs and pAbs, and this was mainly observed with antibodies that strongly labelled the surface of IEs. We demonstrated that this haemolysis was resulted from an immunological interaction between mouse IgMs and a parasite-derived component on the surface of live IEs. Antibodies raised to DBL41 were the most effective in all assays. Of these, three antibodies (pAb B5, mAb B4, mAb G6) and an anti-DBL31 mAb (E7) significantly blocked IE adhesion to purified proteins (ICAM-1 and CD36) under static and flow conditions. These antibodies also blocked parasite adhesion to HUVEC under conditions of blood flow. In a separate work, we characterised the immune response of eight semi-immune serum samples obtained from female adults living in Kilifi County, Kenya. Our results indicated that semi-immune sera specifically recognised five recombinant DBLβICAM-1 domains and a VAR2CSA DBL domain, and recognised the surface of erythrocytes infected by diverse parasite isolates with variable levels of reactivity. Some sera, particularly JA225 and JA235, significantly inhibited IE adhesion to ICAM-1 under both static and flow conditions. To our knowledge, this is the first study to examine the use of PfEMP1ICAM-1-DBLβ domains for the development of mouse mAbs and pAbs that recognise homologous and heterologous parasite isolates and block IE adhesion. However, further work is required to identify the surface ligand(s) involved in interaction with mouse IgM and to investigate the mechanisms of IgM-mediated IE lysis.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QR Microbiology ; QR180 Immunology ; RA0421 Public health. Hygiene. Preventive Medicine