Journal:

Article Title: Glycosylation of the Major Polar Tube Protein of Encephalitozoon hellem , a Microsporidian Parasite That Infects Humans

doi: 10.1128/IAI.72.11.6341-6350.2004

Figure Lengend Snippet: Interaction of EhPTP1 with the lectin ConA. (A) ConA overlay. A DTT-solubilized polar tube preparation was used for SDS-PAGE, transferred to nitrocellulose, and incubated with labeled ConA in the presence and absence of α-methyl-mannopyranoside. ConA bound to a single band (lane 1), and this binding was inhibited by α-methyl-mannopyranoside (lane 2). (B) ConA affinity chromatography. Soluble E. hellem PTPs were loaded onto a ConA-Sepharose column, and specifically retained proteins were eluted with 0.2 M α-methyl-mannopyranoside-0.15 mM NaCl. The flowthrough of the column (e.g., soluble PTPs after loading on the column) and the specific eluate were analyzed by SDS-PAGE followed by immunoblot analysis with rabbit antibody to recEhPTP1. The specific eluate contained EhPTP1 (lane 1), while the flowthrough no longer contained EhPTP1 (lane 2).

Article Snippet: DTT-solubilized PTPs (200 μg) were subjected to ConA affinity chromatography (0.5 ml of ConA gel [EY Laboratories, Inc.]).

Techniques: SDS Page, Incubation, Labeling, Binding Assay, Affinity Chromatography, Western Blot

Journal:

Article Title: Glycosylation of the Major Polar Tube Protein of Encephalitozoon hellem , a Microsporidian Parasite That Infects Humans

doi: 10.1128/IAI.72.11.6341-6350.2004

Figure Lengend Snippet: PTP1 reactivity with ConA is seen in multiple microsporidia. Solubilized (2% DTT) polar tube preparations of five microsporidia species, G. americanus (Ga), E. hellem (Eh), E. cuniculi (Ec), E. intestinalis (Ei), and B. algerae (Ba), were used for a lectin overlay with labeled ConA. ConA reacted with a band in each polar tube preparation that had a size consistent with that seen on SDS-PAGE and immunoblot analysis with antibody to PTP1 (12-15).

Article Snippet: DTT-solubilized PTPs (200 μg) were subjected to ConA affinity chromatography (0.5 ml of ConA gel [EY Laboratories, Inc.]).

Techniques: Labeling, SDS Page, Western Blot

Journal: Cell Proliferation

Article Title: Ability of the polysaccharide chitosan to inhibit proliferation of CD4+ lymphocytes from mucosal inductive sites, in vitro and in vivo

doi: 10.1111/j.1365-2184.2009.00634.x

Figure Lengend Snippet: Effect of chitosan on ConA‐induced T‐cell proliferation in vitro. Mesenteric lymph node (MLN) lymphocytes from normal rats labelled with succinimidyl ester of carboxyfluorescein diacetate (CFSE) were stimulated with 2.5 μg/ml of mitogen in presence or absence of chitosan (1–100 μg/ml) at 37 °C in RPMI 1640 medium supplemented with 10% foetal calf serum in a 95% air/5% CO2 atmosphere for 72 h. After incubation, cells were labelled with anti‐CD4 and 20 000 cells were acquired by flow cytometry. Representative dot plots of CFSE‐labelled cells representing chitosan‐induced inhibition of ConA‐induced mitogenesis are shown. Percentage of daughter cells in the box is included. (a) Unstimulated control and ConA stimulated cells; (b) left: ConA + chitosan; middle: ConA for 24 h and then chitosan for further 72 h; right: chitosan for 24 h and then ConA for further 72 h.

Article Snippet: Compared to the basal condition, the increment was significant either with ConA or chitosan ( P < 0.05) ( ) and highest values were found in cultures with simultaneous addition of both stimuli (ConA + Chi) or pre‐treatment with ConA (ConA24 + Chi) ( P < 0.05).

Techniques: In Vitro, Incubation, Flow Cytometry, Inhibition, Control

Journal: Cell Proliferation

Article Title: Ability of the polysaccharide chitosan to inhibit proliferation of CD4+ lymphocytes from mucosal inductive sites, in vitro and in vivo

doi: 10.1111/j.1365-2184.2009.00634.x

Figure Lengend Snippet: Expression of markers CD25, CD71 and CD95L in lymphocytes stimulated with ConA and chitosan. Mesenteric lymph nodes (MLN) cells (1 × 106 cells/well) cultured for 72 h as described in Fig. 1 were harvested and stained with fluorochrome‐conjugated antibodies against CD4, CD71, CD25 and CD95L as described in the Materials and methods section. (A) Representative dot plots of CD25 and CD71 expression in MLN CD4+ cell cultures with (a) medium (basal); (b) chitosan; (c) ConA; (d) ConA 24 together with chitosan; (e) ConA 24 h and then chitosan; (f) chitosan 24 h and then ConA. (B) Mean of CD25 and CD71expression of 6–8 wells per experimental condition from two similar experiments. (C) Representative histograms of CD95L expression in the same experimental conditions of (A). (D) Mean of CD95L expression of 6–8 wells per experimental condition from two similar experiments. *P < 0.05 vs. basal; #P < 0.05 vs. chitosan or ConA. Other comparisons are indicated with lines in the figure (B and D).

Article Snippet: Compared to the basal condition, the increment was significant either with ConA or chitosan ( P < 0.05) ( ) and highest values were found in cultures with simultaneous addition of both stimuli (ConA + Chi) or pre‐treatment with ConA (ConA24 + Chi) ( P < 0.05).

Techniques: Expressing, Cell Culture, Staining

Journal: Cell Proliferation

Article Title: Ability of the polysaccharide chitosan to inhibit proliferation of CD4+ lymphocytes from mucosal inductive sites, in vitro and in vivo

doi: 10.1111/j.1365-2184.2009.00634.x

Figure Lengend Snippet: Effect of chitosan on ConA‐induced T‐cell proliferation ex vivo. Mesenteric lymph node (MLN) cells from rats fed a single (a) or five doses (b) of diluent (Dil) or of 1–5 mg chitosan (Ch1–5) were labelled with succinimidyl ester of carboxyfluorescein diacetate (CFSE) and stimulated with 2.5 μg/ml ConA as described in the Materials and methods section. After incubation, cells were labelled with anti‐CD4 and 20 000 cells were acquired by flow cytometry. (a) Representative dot plots of CFSE‐labelled cells representing chitosan‐induced inhibition of ConA‐induced mitogenesis are shown. The percentage of daughter cells in the box is included. Bars are means of data from 4–6 rats per treatment. In culture supernatants, the production of INF‐γ was evaluated by ELISA. (b) Representative dot plots of CFSE‐labelled cells representing inhibition of ConA‐induced mitogenesis after sustained administration of chitosan.

Article Snippet: Compared to the basal condition, the increment was significant either with ConA or chitosan ( P < 0.05) ( ) and highest values were found in cultures with simultaneous addition of both stimuli (ConA + Chi) or pre‐treatment with ConA (ConA24 + Chi) ( P < 0.05).

Techniques: Ex Vivo, Incubation, Flow Cytometry, Inhibition, Enzyme-linked Immunosorbent Assay