Validation anti-human FR-β specificity and immunoreactivity.

<p><i>A</i>: Strong staining was appreciated in the placental villous stroma known to express FR-β, validating anti human FR-β binding specificity and immunoreactivity <i>B</i>: Absent staining was shown after staining with the secondary antibody only. Co-expression of the <i>C</i>: anti-CD68 stain and <i>D</i>: FR-β in activated macrophages in a diverticulitis sample validated anti-human FR-β specificity and immunoreactivity.</p

Characteristics breast cancer patients.

<p>Characteristics breast cancer patients.</p

Characteristics ovarian cancer patients.

<p>Characteristics ovarian cancer patients.</p

FR-β expression in representative samples.

<p><i>A</i>: weak expression (staining intensity 1). <i>B</i>: moderate expression (staining intensity 2). <i>C</i>: strong expression (staining intensity 3).</p

<i>Ex vivo</i> histological analysis.

<p>Fluorescent bacterial and tumour cells were imaged using fluorescence microscopy (Nuance). Blue- DAPI (nuclei), Green- U87/GFP, Magenta- <i>B. breve</i>/pCheMC.</p

Bacterial Bioluminescence in the GIT.

<p>The GIT of mice was colonised with either <i>B. breve</i> or <i>E. coli</i> by oral administration of 10⁹ cfu of the relevant strain for three consecutive days. (<b>a</b>) 2D image of athymic mouse 9 days post final feed with <i>B. breve</i> UCC2003/lux. (<b>b</b>) IVIS XR 2D bioluminescence overlayed with X-ray image of mouse 27 days-post feeding with <i>E. coli</i> MG1655/lux. (<b>c</b>) Bacterial counts in specific regions of the GIT 14 days post feeding. Abdominal bioluminescence corresponding to average cfu is also shown (black dots). Data graphed represent the mean ± S.E. (<b>d</b>) Sample isolated images from 3D tomography of mouse from (b). 3D images show a digital mouse atlas of the skeleton to provide anatomical registration. (Movie available in Movie S1). <i>E. coli</i> MG1655 bioluminescence is visible in blue at lower, and white or green at higher levels.</p

Bacterial Growth in Tumours.

<p>(<b>a,b</b>) Bioluminescence from <i>B. breve</i> UCC2003, <i>E. coli</i> MG1655 or <i>S.</i> Typhimurium SL7207 in s.c. LLC tumour bearing mice 11 days following IV delivery. Representative mice are shown. Increase in bacterial bioluminescence in tumours was observed over time (n = 6). Data graphed represent the mean ± S.E. There was no detectable bioluminescence in organs of treated animals, except for <i>S.</i> Typhimurium (<b>c</b>) Representative <i>S.</i> Typhimurium administered mice displaying non-tumour-specific bacterial bioluminescence. Ventral image – Day 3, Dorsal – Day 11. (<b>d</b>) All tumour types examined were colonised by the various strains. (i) <i>B. breve</i>, B16, Day 11 (ii) <i>E. coli</i>, FaDu, Day 7 (iii) <i>B. breve</i>, U87, Day 14.</p

Relationship Between Bacterial Numbers And Bioluminescence.

<p>Viable bacteria in tumours were enumerated by <i>ex vivo</i> bacterial culture from LLC tumours subsequent to BLI at various time-points post IV administration of <i>B. breve</i> or <i>E. coli</i> (n = 6). (<b>a</b>) <b>Bacterial replication in tumours.</b> Increases over time in viable bacterial numbers and bioluminescence. Data graphed represent the mean ± S.E. (<b>b</b>) <b>Correlation between bacterial numbers and bioluminescence in tumours.</b> Direct comparison between photon flux and bacterial colony counts. Log values of bacterial numbers relative to <i>in vivo</i> bioluminescent units are graphed. Correlation between bacterial counts and bacterial bioluminescence signals: R² = 0.84 for <i>B. breve</i>: R² = 0.97 for <i>E. coli</i> which correlates well with previous studies using <i>E. coli</i> MG1655 where R² = 0.94, <i>P</i><0.001 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030940#pone.0030940-Min1" target="_blank">[57]</a>.</p