Inhibitory function of anti-DBPII antibodies in <i>P</i>. <i>vivax</i> individuals against heterologous DBL-TH4 or DBL-TH5 or homologous reference Sal I binding to human erythrocytes measured by COS7 cell binding-inhibition assay

<p>Inhibitory function of anti-DBPII antibodies in <i>P</i>. <i>vivax</i> individuals against heterologous DBL-TH4 or DBL-TH5 or homologous reference Sal I binding to human erythrocytes measured by COS7 cell binding-inhibition assay</p

Polymorphic residues of DBL-TH haplotypes infected in <i>P</i>. <i>vivax</i> patients at the time of enrollment.

<p>Polymorphic residues of DBL-TH haplotypes infected in <i>P</i>. <i>vivax</i> patients at the time of enrollment.</p

Functional inhibition of anti-DBPII antibodies in high responder samples against the panel of DBL-TH variants.

<p>Transfected COS7 cells expressing DBL-TH alleles, DBL-TH1, -TH2, -TH3, -TH4, -TH5, -TH6, -TH7, -TH8, -TH9 and reference Sal I were incubated with 1:100 plasma dilution for 1 hr at 37°C followed by incubation with a 10% suspension of human erythrocytes for 2 hrs. The number of rosettes was compared between wells of transfected cells incubated with plasma relative to wells without plasma (30 fields of view, magnification ×200). The symbols represent mean percent inhibition of two experiments tested in duplicate wells.</p

Antibody recognition of recombinant PvDBPII.

<p>The scatter plot graph shows the anti-DBPII antibody levels in Thai patients compare to naive control as measured by ELISA. <b>(A)</b> Anti-DBPII levels were significantly higher in patients with acute <i>P</i>. <i>vivax</i> than in naive controls, <b>(B)</b> ELISA data classified patients into 3 groups: high responder (HR), low responders (LR) and non-responders (NR). Each dot represents the mean of optical density values in double wells for each sample. The line represents the mean value. Significance was determined by non-parametric analysis using the Mann-Whitney U test. The level of significant was set at <i>P</i> < 0.05.</p

Inhibition efficiency of anti-DBPII antibodies in <i>P</i>. <i>vivax</i> exposed individuals.

<p>A and B: Inhibitory function against a panel DBL-TH haplotypes of vivax patients infected with Sal I strain and had the strongest inhibitory immunity against both heterologous DBL-TH4 and DBL-TH5 strain <b>(A)</b> patient no.9, <b>(B)</b> patient no.15. C, D and E:. Inhibitory function against a panel DBL-TH of vivax patients infected with high polymorphism DBL-TH strain and had the strongest inhibitory immunity against heterologous DBL-TH4 or DBL-TH5 or Sal I strain, <b>(C)</b> patient no.25 infected with DBL-TH2 strain, <b>(D)</b> patient no.37 infected with DBL-TH4 strainand <b>(E)</b> infected with new DBL-TH strain, patient no.30. The transfected COS7 cells expressing Thai DBPII alleles were incubated with plasma and with human erythrocytes. The number of rosettes was compared between wells of transfected cells incubated with antibodies relative to wells without antibodies. Each chart represents the mean of two independent experiments with each dilution tested in triplicate. Error bars represent ± standard deviation. Statistical significance was determined using one-way analysis of variance (ANOVA) and multiple comparison analysis by Bonferroni test.</p

Inhibitory function of anti-DBPII antibodies in <i>P</i>. <i>vivax</i> individuals against heterologous DBL-TH4 and DBL-TH5 or heterologous DBL-TH4, DBL-TH5 and homologous reference Sal I binding to human erythrocytes measured by COS7 cell binding-inhibition assay

<p>Inhibitory function of anti-DBPII antibodies in <i>P</i>. <i>vivax</i> individuals against heterologous DBL-TH4 and DBL-TH5 or heterologous DBL-TH4, DBL-TH5 and homologous reference Sal I binding to human erythrocytes measured by COS7 cell binding-inhibition assay</p

Broad inhibition by high responder samples of erythrocyte binding to DBL-TH haplotypes.

<p>The transfected COS7 cell expressing DBPII reference Sal I or DBL-TH variants were pre-incubated with plasma at dilution 1:500, 1:1000 and 1:2000 for inhibition of DBPII-erythrocyte binding. The charts show the mean inhibition of each DBL-TH variant. Inhibitory function against the panel of DBL-TH variants by (A) HR3 and (B) HR6 samples. Each chart represents the mean of two independent experiments with each dilution tested in triplicate. Error bars represent ± standard deviation. Statistical significance was determined using one-way analysis of variance (ANOVA) and multiple comparison analysis by Bonferroni test.</p

The antibody levels specific to <i>P. vivax</i> antigen.

<p>Graphical display of antibody levels anti-<i>P. vivax</i> shizont protein extract (A) and anti-DBPII (B) in Thai patients (PV), uninfected residents (UR) and naïve controls (NC). Dots represent the mean optical density value in triplicate wells for each sample. Bars represent mean value. Asterisk indicate statistic significant at P<0.05 with non-parametric two independent tests.</p

Naturally acquired anti-DBPII responses and their inhibition efficiency against DBPII binding.

a<p>High responder (HR), Low responder (LR), Non-responder (NR).</p

The synthesized peptides using in antibody purification and testing functional inhibition.

<p>H1, H2 and H3 are target epitopes of naturally acquired inhibitory antibodies <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035769#pone.0035769-Chootong1" target="_blank">[25]</a>. H1.T1, H3.T1, H3.T2, H3.T3 and M3.T1 epitopes are the variant strain among Thai vivax isolates <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035769#pone.0035769-Gosi1" target="_blank">[19]</a>. NI is the target epitopes of non-inhibitory antibodies.</p