biotin conjugated  (Vector Laboratories)

Journal: Journal of Immunology Research

Article Title: CD11c-Specific Deletion Reveals CREB as a Critical Regulator of DC Function during the Germinal Center Response

doi: 10.1155/2018/8947230

Figure Lengend Snippet: Enhanced GC responses in CKO mice: (a) statistical analysis of frozen spleen sections stained with anti-CD3-APC antibody and biotinylated PNA/streptavidin DyLight-488. The number of GCs per mm 2 spleen was counted in WT and CKO mice after immunization with NP-CGG and Alum ( N = 3, unpaired, two-tailed t -test, p = 0.0212). (b) Representative frozen spleen sections from immunized mice stained with anti-CD3-APC antibody (red) for T cells and biotinylated PNA/streptavidin DyLight-488 (green) for GC cells. The slides were imaged with a 10x objective using a fluorescence microscope. (c) ELISA, using plates coated with NIP-BSA for analysis of anti-NP-IgGs. Anti-NP antibodies were detected with anti-mouse IgG. Data indicate the n -fold upregulation of OD values compared to OD-values of unimmunized control mice (mean percentage ± SEM of 5 animals, two-tailed, unpaired t -test, p = 0.0241). (d) Percentages of GL-7 + cells within B220 + cells in spleens of immunized mice determined by flow cytometry (two-tailed, unpaired t -test, p = 0.0018); (e) percentages of CXCR5 + ICOS + cells within CD4 + T cells in spleens of immunized mice determined by flow cytometry; (f) representative dot plot showing T fh cells (as ICOS and CXCR5-expressing cells within CD4 + ) in WT and CKO mice after immunization; (g) representative dot plot showing T fh cells (as PD-1 and CXCR5-expressing cells within CD4 + ) in WT and CKO mice after immunization; (h) percentages of CD11c + cells within CD19 + cells in spleens of immunized mice determined by flow cytometry. For (d, e, and h),each dot represents an individual mouse, and horizontal bars indicate SEM. ∗ p < 0.05 and ∗∗ p < 0.01.

Article Snippet: To detect GCs, sections were stained with biotin conjugated PNA (Vector Laboratories, USA) followed by DyLight488 conjugated streptavidin (Thermo Scientific, USA), and sections were mounted with Fluoromount-G (Southern Biotech, USA).

Techniques: Staining, Two Tailed Test, Fluorescence, Microscopy, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Expressing

Journal: PLoS ONE

Article Title: T Cell and Dendritic Cell Abnormalities Synergize to Expand Pro-Inflammatory T Cell Subsets Leading to Fatal Autoimmunity in B6.NZBc1 Lupus-Prone Mice

doi: 10.1371/journal.pone.0075166

Figure Lengend Snippet: 8-wk-old mice were injected i.p. with OVA or PBS in CFA. The proportions of splenic Tfh cells (CD4 + CD44 hi CD62L lo CXCR5 hi PD1 hi ) and splenic GC B cells (B220 + Fas + PNA hi ) were determined by flow cytometry 2 wks later. (A) Scatterplots showing the proportion of Tfh and GC B cells as a proportion of the CD4 + T cell and B220 + B cell populations, respectively. (B) Scatterplots showing the amount of cytokine produced by OVA-primed splenocytes re-stimulated in-vitro with OVA for 72h. Assays were performed in triplicate and the levels of secreted cytokines measured by ELISA or cytokine bead array (see Methods). Each data point represents the mean of the triplicate with background cytokine production in the absence of antigen subtracted. Horizontal lines indicate the mean of each group examined. Significance levels were determined by one-way ANOVA with Dunns’ post-test. The p values for significant differences between B6 and congenic mouse strains are shown with *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: Biotin-conjugated peanut agglutinin (PNA) was purchased from Sigma-Aldrich and FITC-conjugated anti-CD62L (MEL-14) mAb was purchased from Cedarlane Laboratories (Burlington, ON, Canada).

Techniques: Injection, Flow Cytometry, Produced, In Vitro, Enzyme-linked Immunosorbent Assay