Additional tissues of ARR/ARR clinically affected Cheviot sheep showing PrP^d accumulation.

<p>a) Retina showing diffuse PrP^d accumulation in outer plexiform layer and granular accumulations in inner plexiform layer and retinal ganglion cells. Bar = 50 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. b) Trigeminal ganglion showing granular PrP^d accumulation in satellite cells Bar = 50 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. c) Muscle spindle from ocular muscle showing weak granular PrP^d accumulation in intrafusal muscle fibres (arrows). Bar = 50 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. d) The same muscle spindle as in c) labelled with anti- PrP antibody F99. The same intrafusal muscle fibres as with R145 are labelled (arrows). Bar = 50 µm. IHC with F99 PrP antibody and haematoxylin counterstaining.</p

Western blotting of different brain areas of ARR/ARR clinically affected Cheviot sheep.

<p>a) Western blots of brain tissue with 4 different PrP antibodiesImmunoblots of frontal cortex (FC) thalamus (TH) midbrain (MB) cerebellum (CB) and medulla at the level of the obex (MO) each showed significant accumulation of PrP^res with antibodies P4, L42, and F99. With each of these antibodies the cerebellum showed the greatest concentration of PrP^res. Compared with ARQ/ARQ and VRQ/VRQ scrapie positive control obex samples, the mono-glycosylated band predominates in most of the ARR/ARR samples. The unglycosylated band from all ARR/ARR and positive control samples has a molecular weight of ∼20 to 21 kDa. No labelling or only trace labelling was found when the antibody SAF84 was used. Mol mkr: molecular weight marker (Note: the molecular weight marker produced a very faint signal with L42. To position the weight reference values the blot was digitally overexposed but it is the original, non-saturated blot that is reproduced). b) Graph showing the proportion of di-glycosylated and mono-glycosylated PrP^res for brain from the clinically affected ARR/ARR sheep in comparison with ARQ/ARQ and VRQ/VRQ controls. For each of the brain sites the mono-glycosylated fraction of PrP^res was present in a relatively greater amount than the di-glycosylated fraction when labelled with either P4 or L42 antibodies. In contrast both VRQ/VRQ control and ARQ/ARQ positive controls had a relatively greater amount of di-glycosylated PrP^res. Values for P4 antibody are shown in blue and for L42 in red; frontal cortex, diamonds; thalamus, squares; midbrain, triangles; cerebellum, circles; obex, pentagons; the Suffolk sheep ARQ/ARQ positive control (obex) is outlined in orange and the Cheviot sheep VRQ/VRQ positive control (obex) is outlined in green.</p

Design of the different studies with reference to the source of the inocula used.

<p>In green, inocula used; in blue, source of the inocula; in black use of the inocula for the six different studies on ARR/ARR sheep; in red, use of the same inocula to challenge sheep of susceptible genotypes.</p

PrP^d labelling and vacuolation in the cerebellar cortex of VRQ/VRQ and ARR/ARR sheep.

<p>a) VRQ/VRQ sheep showing diffuse particulate and stellate types of PrP^d accumulation in the cerebellar molecular layer. Vacuolation is relatively subtle. Bar 200 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. b) VRQ/VRQ sheep showing detail of the stellate type of PrP^d accumulation centred on glial cell nuclei in the molecular layer of the cerebellum. Bar 50 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. c) ARR/ARR sheep showing diffuse particulate but not stellate PrP^d accumulation in both molecular and internal granule cell layers and prominent intra-glial types of PrP^d accumulation in the cerebellar molecular layer. Vacuolation is abundant. Bar 200 µm. IHC with R145 PrP antibody and haematoxylin counterstaining. d) ARR/ARR sheep showing detail of the intensity of particulate type of PrP^d accumulation and intense granular PrP^d accumulation associated with glial cell nuclei (intra-microglial type) in the cerebellar molecular layer. Bar 100 µm. IHC with R145 PrP antibody and haematoxylin counterstaining.</p

Design of the different studies.

<p>Age in months for naturally exposed sheep of study 1; months post-infection for all other studies. Sheep pool, pool of 17 scrapie brains (VRQ/VRQ = 6, VRQ/ARQ = 6, ARQ/ARQ = 4, VRQ/ARR = 1).</p><p>Murine strains, sheep dosed with 22 A or 87 V killed at 35 mpi; sheep dosed with ME7 or 79 A killed at 47 mpi. Dose in grams: n/a, not applicable;</p><p>*, four sheep were boosted with 2 g of inoculum by the subcutaneous route (months post-inoculation correspond to those of original challenge).</p

Frequency of accumulation of PrP^d in the LRS tissues of BSE positive sheep.

<p>Frequency of accumulation of PrP^d in the LRS tissues of BSE positive sheep.</p

Minimum Effective Dose of Cattle and Sheep BSE for Oral Sheep Infection - Fig 2

<p>Survival curves of sheep that developed IHC-confirmed BSE according to: a) source and dose of inoculum: dark blue, sheep inocula at 5g dose; light blue, sheep inocula at 0.5/0.05g dose; red, cattle inoculum at 5g dose; orange, sheep inocula at 0.5/0.05g dose. b) recipient breed and dose of inoculum: dark green, Suffolk sheep dosed with 5g; light green, Suffolk sheep dosed with 0.5/0.05g; purple, Romney sheep dosed with 5g; pink, Romney sheep dosed with 0.5/0.05g. Note that in both graphs, most sheep succumbed between ~600 and 1,150 days regardless of dose received and that the few sheep that survived for longer than ~1,150 days did so also regardless of the dose they received.</p

Graphical and schematic representation of the outcome of the experiments in terms of attack rates and survival times.

<p>Attack rates (%), black bars inside white boxes, with actual values next to them. Survival times are indicated as average±SD dpi (age in case of undosed controls). Note that despite the marked differences in AR between the 5g and the 0.5/0.05g doses, STs are very similar with the only significant difference marked as * (for details refer to text).</p

Experimental design.

<p>Experimental design.</p

Additional file 1: Figure S1. of Mitosis in circulating tumor cells stratifies highly aggressive breast carcinomas

Common recognizable Cytologies of CTCs in Mitosis isolated from breast cancer patients with all the “standard” CTC stains from Fig. 1. Figure S2. Common recognizable Cytologies of CTCs in Mitosis isolated from breast cancer patients with all the “standard” CTC stains from Fig. 1. Figure S3. Kaplan-Meier estimates of probabilities of Overall Survival of the patient subpopulations based on receptor status from Fig. 2a (n=33). Figure S4. Box plot of total number of CTCs in each patient versus mitotic CTCs for each patient. Figure S5. CTC counts and Mitotic CTC counts for each patient sample in relation to time of filtration after blood draw. Table S1. Patient subpopulations classified by stage, receptor status and treatment. (PDF 748 kb