Binding of IRBCs to HBMECs in the presence of soluble proteins is expressed as relative binding compared to binding in absence of any protein (No protein). Binding in the presence of serum is expressed as relative binding compared to binding in absence of any serum (No serum). Recombinant gC1qR/HABP1 inhibits binding of IGH-CR14+ (A) to HBMECs in a dose-dependent manner. Recombinant ICAM-1-Fc, CD36-Fc, and BSA have no effect on binding of IGH-CR14+ (A). Anti-gC1qR/HABP1 mouse serum inhibits binding of IGH-CR14+ (B) to HBMECs. Pre-immune mouse serum (PIS), anti-ICAM-1 monoclonal antibody (clone 15.2), and anti-CD36 monoclonal antibody (clone SMΦ) do not have any effect on binding of IGH-CR14+ (B) to HBMECs. The number of IRBCs bound to 100 HBMEC cells was scored in each assay. All data are averages (± standard error) derived from two independent experiments. Each assay was performed in duplicate. Binding of IGH-CR14+ in absence of any protein or serum was in the range of 90–100 IRBCs bound to 100 HBMECs.

Parasite cultures in trophozoite and schizont stages were allowed to form clumps in the presence of PRP. Parasites were stained with acridine orange and the clumping frequency was determined by scoring the frequency of IRBCs found in clumps. Approximately 2,000 to 3,000 IRBCs were scored. Parasite cultures were pre-incubated with recombinant gC1qR/HABP1 or CD36-Fc at different concentrations prior to use in clumping assays to test their ability to inhibit clumping. Clumping frequency in the presence of gC1qR/HABP1 or CD36-Fc is expressed as relative clumping compared to clumping in the presence of PRP alone. Data represent the average (± standard error) of two independent experiments. Each assay was performed in duplicate. (A) IGH-CR14, (B) 3D7, (C) IGH-CR14−, (D) 3D7−, (E) IGH-CR14+, (F) 3D7+.

Binding of IRBCs to HUVECs in the presence of soluble proteins is expressed as relative binding compared to binding in absence of any protein (No protein). Binding in the presence of serum is expressed as relative binding compared to binding in absence of any serum (No sera). Recombinant gC1qR/HABP1 inhibits binding of IGH-CR14+ (A) and 3D7+ (C) to HUVECs in a dose-dependent manner. Recombinant ICAM-1-Fc and BSA have no effect on binding of IGH-CR14+ (A) and 3D7+ (C) to HUVECs. Anti-gC1qR/HABP1 mouse serum inhibits binding of IGH-CR14+ (B) and 3D7+ (D) to HUVECs. Pre-immune mouse serum (PIS), anti-ICAM-1 monoclonal antibody 15.2, and anti-CD36 monoclonal antibody SMΦ do not have any effect on binding of IGH-CR14+ (B) and 3D7+ (D) to HUVECs. The number of IRBCs bound per 100 HUVECs was scored in each assay. All data are averages (± standard error) derived from two independent experiments. Each assay was performed in duplicate. Binding of IGH-CR14+ and 3D7+ IRBCs in absence of any protein or serum was in the range of 150–200 IRBCs bound to 100 HUVECs.

(A) Frequency of platelet-mediated clumping in PRP and PPP. P. falciparum field isolate IGH-CR14, laboratory strain 3D7, and their derivatives, IGH-CR14+ and 3D7+, which bind gC1qR/HABP1, IGH-CR14−, and 3D7−, which bind CD36, and P. falciparum isolate JDP8, which does not bind gC1qR/HABP1 or CD36, were tested for clumping in the presence of PRP and PPP. Parasites in trophozoite and schizont stages were allowed to form clumps in the presence of PRP and PPP. Parasites were stained with acridine orange and the clumping frequency was determined by scoring the frequency of IRBCs found in clumps. Approximately 500 IRBCs were scored for each parasite. The frequency of clumping in the presence of PRP (grey bars) and PPP (black bars) is shown.
(B) Scanning electron micrograph of platelet-mediated clumps formed by IGH-CR14+. Clumps formed by IGH-CR14+ IRBCs in the presence of PRP were analyzed by scanning electron microscopy. Electron micrograph shows several platelets (marked P), which bridge IRBCs in the clumps.
(C) Transmission electron micrograph of platelet-mediated clumps formed by IGH-CR14+. Clumps formed by IGH-CR14+ IRBCs in the presence of PRP were analyzed by transmission electron microscopy. Only IRBCs are present in the clumps. The IRBCs are closely associated with platelets (P) in the clumps.

Parasite cultures in trophozoite and schizont stages were allowed to form clumps in the presence of PRP. Parasites were stained with acridine orange and the clumping frequency was determined by scoring the percentage of IRBCs found in clumps. Approximately 2,000 to 3,000 IRBCs were scored. The ability of anti-gC1qR/HABP1 mouse serum (anti-gC1qR/HABP1) or anti-CD36 mouse monoclonal IgM antibody (anti-CD36, clone SMΦ) to inhibit clump formation was tested by pre-incubating platelets with antibodies prior to use in clumping assays. Pre-immune serum (PIS) from mice immunized with gC1qR/HABP1 and purified mouse IgM were used as controls. Clumping frequency in the presence of anti-gC1qR/HABP1 or anti-CD36 is expressed relative to clumping in the presence of PRP alone. Data represent average (± standard error) of two independent experiments. Each assay was performed in duplicate. (A) IGH-CR14, (B) 3D7, (C) IGH-CR14−, (D) 3D7−, (E) IGH-CR14+, (F) 3D7+.