R&D1801 and RoAb13 recognise overlapping core epitopes in the CCR5 N-terminal domain.

<p>A, R&D1801 or RoAb13 were incubated overnight with competing peptides 1–24 (1 μg/ml) in wells coated with hCCR5_1–31, and binding (shown as OD 405 nm) of antibody was then measured as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128381#pone.0128381.g001" target="_blank">Fig 1a</a> using a rabbit anti-mouse second layer. C—PBS used in place of competitive peptide. B, The peptide sequence of the 24 peptides tested in a. The blue bars shows all peptides which inhibited the binding by more than 50%. C, Diagrammatic representation of the core binding epitope recognized by the two antibodies, defined by peptides which inhibited binding to the whole N-terminal peptide by more than 50%.</p

Immunisation with a chimeric peptide coding a linear CCR5 antibody epitope together with a helper epitope from tetanus toxoid can stimulate antibodies which recognize native CCR5.

<p>A, The sequence of the chimeric peptide used for immunization, showing the CCR5 B cell epitope, the linker sequence and the T cell helper epitope from tetanus toxoid. B, Sera from 5 immunized mice were collected after priming and boosting (see M&M) with the peptide shown in a), and tested in ELISA for binding to hCCR5_1–31. Binding is shown as OD at 405 nm. U: serum from an unimmunized mouse (preimmune sera showed equivalent binding). RoAb13: supernatant from the RoAb13 hybridoma diluted as shown. C, Sera from the same five mice were tested for binding to CCR5 transfectants (red line) or controls at a dilution of 1:50.</p

The sequence and crystals of RoAb13.

<p>A, The protein sequence of RoAb13 (lower sequence) compared to the closest germline sequence for heavy (right) and light (left) chains (as determined by the V-Quest). Conserved residues are shown in red. B, Crystals of RoAb13.</p

Species specificity of RoAb13 and R&D1801.

<p>A, The sequence of the N-terminal extracellular domain of CCR5 from human, rhesus Macaque or mouse CCR5. * identical amino acids—insertion in mouse sequence. B, Flow cytometry showing binding of R&D1801 or RoAb13 to CHO cells transfected with mouse (filled) or human (red line) CCR5. C, Flow cytometry showing binding of R&D1801 or RoAb13 or control to PBMC from rhesus macaques (indirect immunofluorescence, using Goat anti-mouse PE conjugate as second layer). The first panel shows binding of a commercially available PE-conjugated anti-rhesus CCR5 antibody (positive control).</p

Affinity of RoAb13 binding to native cell-surface CCR5 and a peptide coding for the N-terminal extracellular domain of CCR5.

<p>A, RoAb13 was incubated with CCR5-expressing transfected cells or controls at different concentrations and binding measured by indirect immunofluorescence and flow cytometry. The binding was converted to absolute number of bound antibody molecules by using Ig calibration beads as described in M&M and shown in Supporting information <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0128381#pone.0128381.s001" target="_blank">S1 Fig</a>. The graph shows the concentration of bound antibody molecules plotted against the ratio of bound/free antibody (a classical Scatchard plot). The equation showing the relationship between bound and bound/free derived from the law of mass action is shown below the figure, together with the affinity and number of receptors calculated respectively from the slope and intercept of the equation. B, RoAb13 was incubated with different concentrations of hCCR5_1–31N-terminal domain peptide overnight. The remaining amount of free antibody was estimated by binding to hCCR5_1–31 in ELISA and comparison to a standard curve. Bound antibody was calculated as total—free. The resulting bound and bound/free ratio was plotted as in a) and the affinity calculated from the slope.</p

A randomized trial of planned cesarean or vaginal delivery for twin pregnancy

Background: Twin birth is associated with a higher risk of adverse perinatal outcomes than singleton birth. It is unclear whether planned cesarean section results in a lower risk of adverse outcomes than planned vaginal delivery in twin pregnancy.\ud \ud Methods: We randomly assigned women between 32 weeks 0 days and 38 weeks 6 days of gestation with twin pregnancy and with the first twin in the cephalic presentation to planned cesarean section or planned vaginal delivery with cesarean only if indicated. Elective delivery was planned between 37 weeks 5 days and 38 weeks 6 days of gestation. The primary outcome was a composite of fetal or neonatal death or serious neonatal morbidity, with the fetus or infant as the unit of analysis for the statistical comparison.\ud \ud Results: A total of 1398 women (2795 fetuses) were randomly assigned to planned cesarean delivery and 1406 women (2812 fetuses) to planned vaginal delivery. The rate of cesarean delivery was 90.7% in the planned-cesarean-delivery group and 43.8% in the planned-vaginal-delivery group. Women in the planned-cesarean-delivery group delivered earlier than did those in the planned-vaginal-delivery group (mean number of days from randomization to delivery, 12.4 vs. 13.3; P = 0.04). There was no significant difference in the composite primary outcome between the planned-cesarean-delivery group and the planned-vaginal-delivery group (2.2% and 1.9%, respectively; odds ratio with planned cesarean delivery, 1.16; 95% confidence interval, 0.77 to 1.74; P = 0.49).\ud \ud Conclusion: In twin pregnancy between 32 weeks 0 days and 38 weeks 6 days of gestation, with the first twin in the cephalic presentation, planned cesarean delivery did not significantly decrease or increase the risk of fetal or neonatal death or serious neonatal morbidity, as compared with planned vaginal delivery