fitc conjugated mouse anti human cd63  (Bio-Rad)

Journal: Journal of immunological methods

Article Title: Immunofluorescence analysis of human eosinophils.

doi: 10.1016/j.jim.2024.113619

Figure Lengend Snippet: Fig. 3. Colocalization of IL-9 with TfnRcþ recycling endosomes and IL-13 with CD63þ crystalloid granules in PAF-stimulated eosinophils. Human eo sinophils were adhered to glass coverslips were stimulated with PAF for 5 and 60 min. Following fixation and permeabilization, cells were labelled with anti-IL-9 or anti-IL-13 (red) and anti-TfnRc for recycling endosomes or anti-CD63 (green) for crystalloid granules. Nuclei were stained with DAPI (blue). Merged images are shown at right with yellow regions (surface rendered with Imaris software) indicative of colocalization between intracellular cytokines and crystalloid granules. An unstimulated cell stained with IL-13 and CD63 is depicted on the left to showcase morphology. Data is from representative donor. Scale bar represents 5 μm. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Crystalloid granules were immunolabelled using FITC-conjugated mouse anti-human CD63 (Bio-Rad, Hercules, CA) with FITCconjugated mouse IgG1 isotype control (Bio-Rad).

Techniques: Staining, Software

Journal: Journal of immunological methods

Article Title: Immunofluorescence analysis of human eosinophils.

doi: 10.1016/j.jim.2024.113619

Figure Lengend Snippet: Fig. 4. Object-based colocalization of IL-13þ and CD63þ granules in human eosinophils. Within 2 μm of the cell volumes, intracellular IL-13+ and CD63+

Article Snippet: Crystalloid granules were immunolabelled using FITC-conjugated mouse anti-human CD63 (Bio-Rad, Hercules, CA) with FITCconjugated mouse IgG1 isotype control (Bio-Rad).

Techniques:

Journal: JCI Insight

Article Title: Sialylation regulates neutrophil transepithelial migration, CD11b/CD18 activation, and intestinal mucosal inflammatory function

doi: 10.1172/jci.insight.167151

Figure Lengend Snippet: ( A and B ) Human PMN were exposed to 5 mM Sia, 5 mM Gal, 5 mM 2-DN, or 5 mM KDO for 30 minutes at 37°C, followed by stimulation with 1.25 μM LaB and 5 μM fMLF to induce degranulation before assessment of surface expression of CD66b and CD63 by flow cytometry. Data shown are fold-change in mean fluorescence intensity (MFI) comparing treatment with sialidase inhibitors against relevant control (Gal or KDO). Data are expressed as mean ± SEM and were analyzed by 1-way ANOVA followed by Bonferroni post hoc testing ( n = 4–6 PMN donors, *** P < 0.001, **** P < 0.0001). ( C and D ) Murine PMN were exposed to 5 mM Sia, 5 mM Gal, 5 mM 2-DN, or 5 mM KDO for 30 minutes at 37°C followed by stimulation with 1.25 μM LaB and 10 μM fMLF to induce degranulation before assessment of surface expression of CD66b and CD63 by flow cytometry. Data shown are fold-change in mean fluorescence intensity (MFI) comparing treatment with sialidase inhibitors against relevant control (Gal or KDO). Data are expressed as mean ± SEM and were analyzed by 1-way ANOVA followed by Bonferroni post hoc testing for PMN isolated from 3–5 mice (** P < 0.01, *** P < 0.001). ( E ) Human PMN incubated with 5 mM Gal, 5 mM Sia, 5 mM 2-DN, or 5 mM KDO were exposed to 100 μM cytochrome C. Reduction of cytochrome C in response to 500 nM fMLF was measured by quantifying changes in absorbance at 550 nm at 2, 5, 10, 15, 20, and 60 minutes. Data are fold change in absorbance relative to time 0, are expressed as mean ± SEM, and were analyzed by 1-way ANOVA followed by Bonferroni post hoc testing ( n = 3 independent human PMN donors, * P < 0.05, ** P < 0.01, *** P < 0.001). ( F ) Murine PMN incubated with 5 mM Gal, 5 mM Sia, 5 mM 2-DN, or 5 mM KDO were exposed to 100 μM cytochrome C. Reduction of cytochrome C in response to 1 μM fMLF was measured by quantifying changes in absorbance at 550 nm at 2, 5, 10, 15, 20, and 60 minutes. Data are fold change in absorbance relative to time 0; data are expressed as mean ± SEM and were analyzed by 1-way ANOVA followed by Bonferroni post hoc testing ( n = 3 mice, * P < 0.05, ** P < 0.01, *** P < 0.001).

Article Snippet: Mouse anti–human FITC–conjugated anti-CD63 (catalog 550759) and anti-CD66b (catalog 561927) mAbs were purchased from BD Bioscience Rat anti–mouse PE-conjugated CD63 (catalog 12063182) and anti-CD15 (catalog MA1-022) mAbs were purchased from Thermo Fisher Scientific.

Techniques: Expressing, Flow Cytometry, Fluorescence, Isolation, Incubation