Immunocytochemistry.

<p>Fluorescence microscopy-based detection of PSMA-positive prostate cancer cell line with muJ591:MIRB complex: Secondary antibody staining conjugated to AlexaFluor-488 shows the binding of the J591 antibody in (A) LNCaP and whereas (B) DU145 control cells have no binding; and (C) Rhodamine-B fluorophore was observed on LNCaP cells (PSMA-positive) and not on (D) DU145 cells (PSMA-negative).</p

T2 relaxation time of treated cells.

<p>A. T2 relaxation time as a function of the iron concentration for PSMA-positive (LNCaP) and PSMA-negative (DU145) live prostate cancer cells treated with the antibody:MIRB conjugates (n = 3 for each concentration); B. Corresponding MR parametric maps of T2 relaxation times (color) and intensity imaging (TE = 5.8 ms) of samples treated with different antibody:MIRB concentrations (0.28, 0.14, 0.07, 0.04, 0.02, 0.01 µg/µL protein concentration equivalent to 0.2, 0.1, 0.05, 0.03, 0.012, 0.006 µg/µL in iron).</p

T2 relaxation time for different seeded counts of LNCaP treated cells.

<p>A. T2 relaxation time as a function of the originally seeded LNCaP cells (n = 3); B. Corresponding MR parametric maps of T2 relaxation times (color) and intensity imaging (TE = 5.8 ms) of samples. All cells were treated with muJ591:MIRB at 0.28 µg/µL protein concentration equivalent to 0.2 µg/µL in iron.</p

Geometric mean and median of 10,000 events obtained by flow cytometry for different cell lines and treatment with the antibody and antibody:MIRB conjugates.

<p>Geometric mean and median of 10,000 events obtained by flow cytometry for different cell lines and treatment with the antibody and antibody:MIRB conjugates.</p

muJ591:MIRB characterization.

<p>A. Scanning Electron Micrograph (SEM) of immobilized muJ591:MIRB complexes and B. Energy Dispersive X-ray (EDX) mapping for Fe, C, O and N superimposed on the SEM. The presence of MIRB nanoparticles can be observed as iron signal (solid arrow) and the antibody as organic matter signal (dashed arrow). The large white structures are salts from the dried buffer.</p

Flow cytometry.

<p>Specificity and uptake of muJ591:MIRB, muIgG:MIRB, muJ591 and muIgG of prostate cancer cells. A. LNCaP cells presented a shift that showed uptake of the muJ591 and of the conjugate muJ591:MIRB and no uptake for the controls muIgG antibody or muIgG:MIRB antibody conjugate. B. Control DU145 cells did not show uptake.</p

T2 relaxation time of conjugates.

<p>A. T2 relaxation time as a function of the iron concentration for the conjugates and free MIRB nanoparticles (n = 3 for each iron concentration); B. Corresponding MR parametric maps of T2 relaxation times (color) and intensity imaging (TE = 5.8 ms) of samples at different iron concentrations for MIRB alone, muIgG:MIRB and muJ591:MIRB conjugates.</p

Fluorimetric changes.

<p>Effect of antibody conjugation on the relative fluorescence intensity of Molday ION Rhodamine-B complex (n = 3).</p

Models for cohesion and cohesion-mediated chromosome segregation.

<p>(A) SeqA-dependent precatenane removal. Positive supercoils migrate behind the replisome, entwining newly replicated sister regions. Resolution of precatenanes by Topo IV (green) is delayed by SeqA (red), which binds to hemimethylated DNA tracts behind the fork. Five to ten minutes after fork passage, DNA is remethylated by Dam (blue), releasing SeqA, and allowing Topo IV to resolve inter-sister links. SeqA may inhibit Topo IV by restraining supercoils, which transforms Topo IV-reactive hooked juxtaposition crossings (lower left brackets) to Topo IV-unreactive unhooked crossings (lower right brackets). (B) Simultaneous release of cohesion along right-arm snap regions (red) promotes abrupt sister separation and results in individualized daughter nucleoids (middle). Premature cohesion loss results in poor sister individualization (top). Deficient removal of cohesion results in late/unfinished sister separation (bottom).</p

Genetic interactions between Topo IV and SeqA.

<p>(A) <i>ΔseqA</i> partially suppresses the temperature sensitivity of <i>parE10</i>. <i>parE10</i> single and Δ<i>seqA parE10</i> double mutant cells were grown to exponential phase at 30°C, equal numbers of cells were plated and incubated at the indicated temperature for 24 h and colonies were photographed. (B) Quantification of colony formation data. Wild-type, Δ<i>seqA</i> and <i>parE10</i> single and double mutants were plated and grown as described above, and the number of colonies relative to WT grown at 30°C were plotted (2 independent experiments ±1 SD). (C) Cohesion relationship between SeqA and Topo IV. Cohesion values were determined at <i>gln</i> and <i>dnaB</i> in cells carrying loss-of-function alleles or overexpression constructs of <i>seqA</i> and <i>parE</i>. Overexpression (OE) was achieved by transformation with low-level expression plasmids (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003673#s4" target="_blank">Materials and Methods</a>). Vector plasmids showed wild-type cohesion levels at <i>gln</i> and <i>dnaB</i> (1.31±0.07 and 1.05±0.04 copies/focus, respectively).</p