Journal: PLoS Pathogens

Article Title: Prion Uptake in the Gut: Identification of the First Uptake and Replication Sites

doi: 10.1371/journal.ppat.1002449

Figure Lengend Snippet: Peyer's patches were collected from wild-type 29/Ola mice 1 day after infection with ME7 prions. (A) Cellular organization of a Peyer's patch showing the LAMP1-positive compartments at the FAE and SED. (B) FAE enterocytes have large LAMP1-positive endosomes compared to enterocytes in the villi. (C and D) UEA-1 positive (red) M cells within the FAE. Note in (D) the large apical LAMP1+ endosomes are not present in M cells, but only in adjacent FAE enterocytes (arrowheads). N depicts the approximate location of nucleus in the epithelium (E) UEA-1 label is present only on FAE, whereas the neighbouring villus is negative. IgG (green) presents the endogenous mouse IgGs. (F and G) Cryo-immuno EM reveals that M cells in the FAE differ from enterocytes by having irregular microvilli that are UEA-1–positive (F, G). The diameter of the UEA-1–specific gold particles in (F) and (G) is 15 nm. Note in (F) the arrow points to a PrP-positive early endosomal vesicle in the UEA-1 negative enterocyte, whereas little PrP uptake is detectable in the UEA-1 positive M cell. PrP was detected with PrP-specific 6H4 monoclonal antibody directly conjugated to UltraSmall gold (PrP-usg) and visualized by silver enhancement. A larger high resolution version of is provided as a supplement . SED, subepithelial dome; GC, germinal centre; V, villus; M, M cells; E, enterocytes. Scale bars: (A) 80 µm; (B) 15 µm; (C) 10 µm; (D) 15 µm; (E) 20 µm; (F) 200 nm and (G) 200 nm.

Article Snippet: M cells were identified with biotinylated Ulex europaeus (UEA-1) lectin (L8262; Sigma).

Techniques: Infection

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fc? Receptor Upregulation Is Associated With Immune Complex Inflammation in the Mouse Retina and Early Age-Related Macular Degeneration

doi: 10.1167/iovs.13-11821

Figure Lengend Snippet: Deposition of IgG and C1q and complement activation are observed in the choriocapillaris of patients with early AMD. (a) Immunohistochemical detection of IgG (green) and U. europaeus agglutinin I (red) (n = 9–11 per group) revealed IgG deposition in close proximity to choriocapillaris in both control and early AMD eyes. (b) Immunohistochemical detection of IgG (green) and C1q (red) (n = 9 per group) in aged-matched controls and patients with early AMD showed colocalization of IgG and C1q (arrows) in blood vessel lumens, indicating formation of immune complexes but not in drusen (star) (c) Immunohistochemical detection of IgG (green) and MAC (C5b-9, red) in aged-matched controls and patients with early AMD (n = 9 per group) revealed MAC and IgG colocalization around the choriocapillaris (arrowheads) and in drusen (star) in early AMD (inset). Note the intense autofluorescence of the RPE due to lipofuscin autofluorescence. CC, choriocapillaris. Scale bars: 50 μm and 25 μm (inset).

Article Snippet: The primary antibodies used were the following: FITC-labeled donkey F(ab′) 2 fragment anti-human IgG (Jackson ImmunoResearch Laboratories), polyclonal rabbit anti-C1q (Dako, Cambridge, UK), monoclonal mouse anti-MAC-C5b-9 (aE11; Dako), monoclonal mouse anti-human CD45 (HI30; BD Biosciences, Oxford, UK), monoclonal rabbit anti-FcγRIIb (EP888Y; Abcam), monoclonal rabbit anti-FcγRIIa (EPR6658; Epitomics–an Abcam Company, Burlingame, CA), monoclonal mouse anti-FcγRIII (2h7; Abcam), polyclonal goat anti-FcγRI (R&D Systems), and biotinylated Ullex europaeus agglutinin I (UEA1; Vector Laboratories).

Techniques: Activation Assay, Immunohistochemical staining

Journal: Arthritis Research & Therapy

Article Title: Fucosyltransferase 1 mediates angiogenesis, cell adhesion and rheumatoid arthritis synovial tissue fibroblast proliferation

doi: 10.1186/ar4456

Figure Lengend Snippet: α(1,2)-linked fucosylated proteins are expressed in rheumatoid arthritis (RA). (A) RA synovial tissue (ST) homogenates contained more α(1,2)-linked fucosylated proteins than did osteoarthritis (OA) or normal (NL) STs (normalized to total protein concentration). Results are expressed as a ratio of the amount of fucosylated BSA/total proteins in the ST homogenates using BSA as a standard. (B) Photographs of RA ST. The far left panel shows staining with mouse anti-collagen 1. The middle panel shows staining with Ulex Europeaus Agglutinin 1 lectin (UEA-1) and goat anti-UEA-1. The right panel shows merging of the left panel and middle panel. Yellow indicates α(1,2)-linked fucosylated proteins associated with RA ST fibroblasts and the blue indicates DAPI staining of the tissue (original magnification 100×). (C) α(1,2)-linked fucosylated proteins in RA synovial fibroblast-conditioned medium were significantly higher than in OA and NL synovial fibroblast-conditioned medium. (D) α(1,2)-linked fucosylated proteins in RA synovial fibroblast cell lysates were significantly higher than in NL synovial fibroblast cultures: 2′fucosyllactose-bovine serum albumin (2′FL-BSA) was used as a standard.

Article Snippet: UEA-1, 2 μg/ml (Vector laboratories Inc, Burlingame, CA, USA) was added for 90 minutes followed by 10 μg/ml biotinylated goat anti-UEA-1 antibody (Vector laboratories Inc) for 60 minutes, then streptavidin-horseradish peroxidase (HRP) for 30 minutes.

Techniques: Protein Concentration, Staining