staining for muc1  (Santa Cruz Biotechnology)

Journal: Frontiers in Immunology

Article Title: Global Inhibition of DC Priming Capacity in the Spleen of Self-Antigen Vaccinated Mice Requires IL-10

doi: 10.3389/fimmu.2014.00059

Figure Lengend Snippet: Splenic DC activation is suppressed as early as 4–8 h post-vaccination with a self-, but not a foreign-antigen and correlated with early IL-10 production in the spleens of these animals . WT (squares) and MUC1.Tg mice (triangles) were vaccinated with MUC1p plus Poly-ICLC via tail vein. Spleens were removed at indicated hours post-vaccination and total splenic mRNA levels of trypsin 1 (A) , carboxypeptidase B1 (CPB1) (B) , and IL-10 (C) were determined relative to the control gene HPRT. Values shown represent expression relative to the baseline expression in mice of that genotype (WT and MUC1.Tg) at 0 h post-vaccination. Data are representative of three pooled mice were group per time point shown. Data points show mean ± SEM of three technical replicates and are representative of two independent experiments.

Article Snippet: These DC were put into culture with bead isolated (CD4 T cell Isolation Kit II, Miltenyi) CFSE stained MUC1 specific VFT CD4 T cells ( ) at a ratio of 1 DC to 5 VFT cells in complete DMEM.

Techniques: Activation Assay, Control, Expressing

Journal: Frontiers in Immunology

Article Title: Global Inhibition of DC Priming Capacity in the Spleen of Self-Antigen Vaccinated Mice Requires IL-10

doi: 10.3389/fimmu.2014.00059

Figure Lengend Snippet: Dendritic cell from spleens of mice vaccinated with self-antigen have higher levels of phosphorylated STAT3 after IL-10 treatment than DC from spleens of mice vaccinated with foreign-antigen . WT (solid line) and MUC1.Tg (dashed line) mice were vaccinated with MUC1p via the tail vein. Twenty-four hours post-vaccination splenocytes were removed and treated with 30 ng/mL of IL-10 for 20 min. Following incubation, cells were fixed and phospho-STAT3 expression in CD11c+NK1.1− splenocytes was analyzed via phoshpoflow. (A) A representative flow plot is shown. The shaded histogram represents the fluorescence level when cells are treated with standard surface markers and an isotype-matched control instead of the phosphospecific antibody. pSTAT3 positivity (B) and MFI (C) were analyzed. In (B) symbols correspond to individual animals and are representative of two independent experiments. (C) Values shown have been normalized to the expression level of the control group (WT) in order to allow for pooling of data from separate experiments run on multiple days. Bars are representative of nine mice from two combined experiments and show the mean ± SEM. *Indicates a p -value of <0.05.

Article Snippet: These DC were put into culture with bead isolated (CD4 T cell Isolation Kit II, Miltenyi) CFSE stained MUC1 specific VFT CD4 T cells ( ) at a ratio of 1 DC to 5 VFT cells in complete DMEM.

Techniques: Incubation, Expressing, Fluorescence, Control

Journal: Frontiers in Immunology

Article Title: Global Inhibition of DC Priming Capacity in the Spleen of Self-Antigen Vaccinated Mice Requires IL-10

doi: 10.3389/fimmu.2014.00059

Figure Lengend Snippet: Pretreatment with an antibody against the IL-10 receptor increases the level of costimulatory molecule expression on DC in the spleens of self-antigen vaccinated mice . MUC1.Tg mice were pretreated with an antibody against the IL-10 receptor (IL-10R, solid lines) or were given a non-specific isotype control (iso, dashed lines). One to two days later they were vaccinated as in Figure and 24 h post-vaccination, splenocytes were removed and analyzed via flow cytometry. The expression level of CD40 (A) , CD86 (B) , and CD80 (C) on splenic DC (CD11C+, MHC Class II+) was determined. Shaded histograms represent fluorescence in samples stained with isotype alone. Bar graph values shown have been normalized to the expression level of the control group (iso) in order to allow for pooling of data from separate experiments run on multiple days. (A,C) Data are combined from two independent experiments and representative of six mice. (B) Data are combined from three independent experiments and are representative of 10 mice. Bars represent mean ± SEM. p -Values are as stated unless designated by a *, which indicates a p -value of <0.05.

Article Snippet: These DC were put into culture with bead isolated (CD4 T cell Isolation Kit II, Miltenyi) CFSE stained MUC1 specific VFT CD4 T cells ( ) at a ratio of 1 DC to 5 VFT cells in complete DMEM.

Techniques: Expressing, Control, Flow Cytometry, Fluorescence, Staining

Journal: Frontiers in Immunology

Article Title: Global Inhibition of DC Priming Capacity in the Spleen of Self-Antigen Vaccinated Mice Requires IL-10

doi: 10.3389/fimmu.2014.00059

Figure Lengend Snippet: Blocking of the IL-10 receptor prior to intravenous MUC1 peptide vaccination increases the ability of splenic DC from MUC1.Tg mice to stimulate MUC1 specific CD4 T cells ex vivo . MUC1.tg mice were treated as in Figure . Twenty-four hours post MUC1 vaccination, splenocytes from three to four mice per treatment group were pooled and bead isolated DC from these pooled splenocytes were put into culture with CFSE labeled MUC1 specific CD4 T cells (VFT cells) at a ratio 1DC:5VFT. Twenty-four hours after the start of culture, half of the culture media was removed and the concentration of IL-2 was measured by ELISA (A) . Cultures were allowed to incubate three more days for a total of four and VFT proliferation was analyzed by CFSE dilution (B,C) . (B) Bars represent the mean percentage of CD3+CD4+ T cells that had proliferated at 4 days of three technical replicates ±SEM. (C) A representative flow plot is shown. Data are representative of two to three independent experiments. *Indicates a p -value of <0.05.

Article Snippet: These DC were put into culture with bead isolated (CD4 T cell Isolation Kit II, Miltenyi) CFSE stained MUC1 specific VFT CD4 T cells ( ) at a ratio of 1 DC to 5 VFT cells in complete DMEM.

Techniques: Blocking Assay, Ex Vivo, Isolation, Labeling, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: Global Inhibition of DC Priming Capacity in the Spleen of Self-Antigen Vaccinated Mice Requires IL-10

doi: 10.3389/fimmu.2014.00059

Figure Lengend Snippet: Treatment with anti-IL-10R antibody at the time of vaccination increases the number of MUC1p specific, IFNγ+ CD4 T cells without an effect on CD8 T cells . WT and MUC1.Tg mice were pretreated with an antibody against the IL-10 receptor (IL-10R, black bars) or a non-specific isotype control (iso, gray bars). One to two days following antibody treatment, mice were vaccinated with DC loaded with MUC1p. Seven to nine days post-vaccination, spleens were removed and bead isolated CD4 (A,C) and CD8 T cells (B,D) were cultured with MUC1p loaded bone marrow derived DC overnight and analyzed by ELISPOT (A,B) , or were cultured for 6–8 h in the presence of brefeldin-A and analyzed by intracellular flow cytometry (C,D) . (A) Data are combined from two independent experiments with each spot indicating an individual animal. Data are representative of three independent experiments. (B) Bars indicate the average of three technical replicates pooled from three individual animals per group. Data are representative of two independent experiments. (C,D) Values shown are normalized to the response of mice of that genotype (WT versus MUC1.Tg) given the control treatment (iso). Data are combined from two independent experiments and are representative of five to six mice per group. Bars represent mean ± SEM. *Indicates a p -value of <0.05; **indicates a p -value of <0.005.

Article Snippet: These DC were put into culture with bead isolated (CD4 T cell Isolation Kit II, Miltenyi) CFSE stained MUC1 specific VFT CD4 T cells ( ) at a ratio of 1 DC to 5 VFT cells in complete DMEM.

Techniques: Control, Isolation, Cell Culture, Derivative Assay, Enzyme-linked Immunospot, Flow Cytometry

Journal: Scientific reports

Article Title: De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering.

doi: 10.1038/s41598-024-59471-z

Figure Lengend Snippet: Figure 2. Characterization of eMSCs (a) and eECs (b). (A) The outgrowth of the mix population from the explanted tissue under MSC medium condition. (B) The eMSCs had a spindle-shaped morphology, and (D) positive Vimentin staining indicated the presence of this intermediate filament protein (red: nuclei in blue). (C) MSC-specific markers (CD29, 40, 90) were demonstrated by gene expression analysis. (E) As indicators of osteogenic potential, eMSCs exhibited positive ALP staining, and (F) positive alizarin red staining for extracellular calcium deposits under osteogenic culture conditions, confirming differentiation into osteogenic lineages. (G) Oil Red O staining emphasized the presence of intracellular lipid droplets, confirming adipogenic differentiation capability of the eMSCs. The characterization of eECs (b) revealed (H,K) The outgrowth of the epithelial cells from the explanted tissue under FBS-EC and KSR-EC medium condition respectively (I,N) a polygonal cell shape in both EC media tested (FBS-EC and KSR-EC, respectively). (J,O) Positive Pan Cytokeratin staining of the cytoskeleton was evident in eECs in both media. In FBS-EC media, eEC purity, determined by flow cytometry, was 94.78% (K), whereas in KSR media, purity reached 97.70% (P). (L,Q) PCR validated the gene expression for the eEC-specific marker Muc1 in eECs derived from both EC media. (Uncropped conventional PCR gels were shown in the Supplementary Uncropped Fig. 1A–F for eMSCs and Figure for eECs).

Article Snippet: Additionally, the secretory function of the endometrial glandlike structures was assessed by staining for MUC1 (sc-7313, Santa Cruz Biotechnology), with a protocol adapted from Piña et al.31.

Techniques: Staining, Gene Expression, Flow Cytometry, Marker, Derivative Assay

Journal: Scientific reports

Article Title: De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering.

doi: 10.1038/s41598-024-59471-z

Figure Lengend Snippet: Figure 4. Morphology of eECs from passage 6 in; (A) KSR-EC medium, and (B) FBS-EC medium. (C) Muc1 gene expression was not detected in eECs cultured in KSR-EC medium. (D) In contrast, eECs cultured in FBS-EC medium did show Muc1 expression. Scale bar: 300 µm. (Uncropped conventional PCR gels were shown in the Supplementary Fig. 2).

Article Snippet: Additionally, the secretory function of the endometrial glandlike structures was assessed by staining for MUC1 (sc-7313, Santa Cruz Biotechnology), with a protocol adapted from Piña et al.31.

Techniques: Gene Expression, Cell Culture, Expressing

Journal: Scientific reports

Article Title: De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering.

doi: 10.1038/s41598-024-59471-z

Figure Lengend Snippet: Figure 6. (A) Expression of the FOXA2 gene in in vivo endometrial tissue and in the in vitro 3D-ET. FOXA2 protein was localized to the nucleus of the in vitro gland-like structures. (B) Muc1 gene expression in in vitro 3D-ET and (C) in vivo tissue with MUC1 protein expression in the cytoplasm of the cells lining the gland. (Uncropped conventional PCR gels were shown in the Supplementary Fig. 5).

Article Snippet: Additionally, the secretory function of the endometrial glandlike structures was assessed by staining for MUC1 (sc-7313, Santa Cruz Biotechnology), with a protocol adapted from Piña et al.31.

Techniques: Expressing, In Vivo, In Vitro, Gene Expression

Journal: Breast Cancer Research : BCR

Article Title: Polarity gene alterations in pure invasive micropapillary carcinomas of the breast

doi: 10.1186/bcr3653

Figure Lengend Snippet: Phenotypic and genomic characterisation of invasive micropapillary carcinoma. (A) and (B) Histological samples of an invasive micropapillary carcinoma (IMPC) of the breast composed of cell clusters surrounded by empty spaces and displaying an inside-out growth pattern, as highlighted by MUC1 staining. (C) Frequency plots of gains and losses are displayed from chromosome 1pter on the left to chromosome Xq on the right. Alternating grey and white bands indicate chromosome boundaries. Dashed blue line represent 40% frequencies, − for losses and + for gains, respectively. ER, Oestrogen receptor; IDC-NST, Invasive ductal carcinoma of no special type.

Article Snippet: IMPC cases were confirmed on the basis of inside-out MUC1 staining (cancer antigen 15-3 (CA 15-3), monoclonal antibody, clone DF3; AbCys, Paris, France) at the inverted apical pole [ ].

Techniques: Staining