af680- albumin  (Thermo Fisher)

Journal: Microcirculation (New York, N.y. : 1994)

Article Title: Regulatory T Cells Control Vascular Adhesion Molecule Expression in Skin Under Inflammatory and Homeostatic Conditions

doi: 10.1111/micc.70017

Figure Lengend Snippet: CHS induces upregulation of E‐selectin and ICAM‐1 in the dermal microvasculature. Adhesion molecule expression in the dermal microvasculature in a two‐challenge model of CHS was assessed in wild‐type (WT) C57BL/6 mice 4 h after the second challenge, via in vivo labelling with fluorochrome‐conjugated antibodies. Tissues underwent intravascular staining followed by exsanguination, fixation, clearing and imaging via multiphoton microscopy, with samples collected from CHS skin and untreated skin from the same mice. (A, B) Representative images of staining for CD31 (vasculature), E‐selectin and ICAM‐1 (A) or CD31, P‐selectin and VCAM‐1 (B) either in untreated skin (top row in A, B) or during CHS (bottom row in A, B). Images are maximum intensity projections through the skin, taken from the epithelial surface to ~400 μm depth in inflamed samples. Right‐hand panels show merged images. Scale bars = either 200 μm (A: WT untreated; B: WT CHS) or 150 μm (A: WT CHS; (B) WT Untreated). (C) Isotype control antibody staining in the dermal microvasculature during two challenge CHS. Mice underwent the two‐challenge model of CHS and received AF594‐anti‐CD31, along with AF680‐mouse IgG1κ (isotype control for RME‐1/anti‐E‐selectin) and AF488‐rat IgG2b (isotype control for YN1.1.7.4/anti‐ICAM‐1), and the skin was cleared and imaged as for the other samples. Images are shown for staining for CD31 (red), and the AF680‐conjugated (cyan) and AF488‐conjugated (green) isotype control antibodies. Scale bar = 150 μm. (D, E) Quantitation of adhesion molecule expression in untreated (UT) or CHS skin for E‐selectin and P‐selectin (D) or ICAM‐1 and VCAM‐1 (E). Data from samples treated with isotype control antibodies conjugated with AF680 (D) or AF488 (E) are also shown. Data are shown as mean ± sem with data for individual mice also shown. N = 3/group for adhesion molecules and 4/group for isotype controls. * p < 0.05 for comparison of UT and CHS (in the same animal) via paired t ‐test for each adhesion molecule.

Article Snippet: To enable visualization of the vasculature, AF680‐albumin (25 μg/mouse, Thermo Fisher Scientific) was administered i.v.

Techniques: Expressing, In Vivo, Staining, Imaging, Microscopy, Control, Quantitation Assay, Comparison

Journal: Microcirculation (New York, N.y. : 1994)

Article Title: Regulatory T Cells Control Vascular Adhesion Molecule Expression in Skin Under Inflammatory and Homeostatic Conditions

doi: 10.1111/micc.70017

Figure Lengend Snippet: Treg depletion promotes increased expression of adhesion molecules in the dermal microvasculature during CHS. Adhesion molecule expression in the dermal microvasculature during two‐challenge CHS was assessed in wild‐type (WT) mice and Foxp3 DTR mice treated with diphtheria toxin (DT) to deplete Tregs, commencing 24 h after the initial oxazolone challenge. Adhesion molecule expression was assessed 4 h after the second challenge as described in Figure . (A, B) Representative images of staining for CD31 (vasculature), E‐selectin and ICAM‐1 (A) or CD31, P‐selectin and VCAM‐1 (B) in DT‐treated wild‐type mice (WT + CHS + DT) or DT‐treated Foxp3 DTR mice ( Foxp3 DTR + CHS + DT). Right‐hand panels show merged images. Scale bars = 100 μm. (C–F) Quantitation of adhesion molecule expression in DT‐treated wild‐type and Foxp3 DTR mice undergoing CHS. Data are shown for E‐selectin (C), ICAM‐1 (D) P‐selectin (E) and VCAM‐1 (F). Data are shown as mean ± sem with data for individual mice also shown. N = 5 for WT (E‐selectin/ICAM‐1), 8 for Foxp3 DTR (E‐selectin/ICAM‐1), 7 for WT (P‐selectin/VCAM‐1) and 6 for Foxp3 DTR (P‐selectin/VCAM‐1). * p < 0.05 for comparisons shown via unpaired t ‐test.

Article Snippet: To enable visualization of the vasculature, AF680‐albumin (25 μg/mouse, Thermo Fisher Scientific) was administered i.v.

Techniques: Expressing, Staining, Quantitation Assay

Journal: Microcirculation (New York, N.y. : 1994)

Article Title: Regulatory T Cells Control Vascular Adhesion Molecule Expression in Skin Under Inflammatory and Homeostatic Conditions

doi: 10.1111/micc.70017

Figure Lengend Snippet: Treg depletion promotes increased expression of E‐selectin and ICAM‐1 in the uninflamed dermal microvasculature. Adhesion molecule expression in the dermal microvasculature of untreated skin was compared in Treg‐intact wild‐type mice and Treg‐depleted Foxp3 DTR mice 28 h after initiation of Treg depletion. Adhesion molecule expression was assessed as described in Figure . (A) Representative images of staining for CD31 (vasculature), E‐selectin, and ICAM‐1 in untreated skin of DT‐treated wild‐type mice (UT WT + DT) or DT‐treated Foxp3 DTR mice (UT Foxp3 DTR + DT). Right‐hand panels show merged images. Scale bars = 100 μm (UT WT + DT) or 150 μm (UT Foxp3 DTR + DT). (B–E) Quantitation of adhesion molecule expression in untreated skin of DT‐treated wild‐type (WT) and Foxp3 DTR mice. Data are shown for E‐selectin (B), ICAM‐1 (C) P‐selectin (D) and VCAM‐1 (E). Data are shown as mean ± sem with data for individual mice also shown. N = 6 for WT (E‐selectin/ICAM‐1), 8 for Foxp3 DTR (E‐selectin/ICAM‐1), 7 for WT (P‐selectin/VCAM‐1) and 7 for Foxp3 DTR (P‐selectin/VCAM‐1). * p < 0.05 for comparisons shown via unpaired t ‐test. ns, not significant.

Article Snippet: To enable visualization of the vasculature, AF680‐albumin (25 μg/mouse, Thermo Fisher Scientific) was administered i.v.

Techniques: Expressing, Staining, Quantitation Assay