Journal:

Article Title: Upregulation of CD40 Expression on Endothelial Cells Infected with Human Cytomegalovirus

doi: 10.1128/JVI.76.24.12803-12812.2002

Figure Lengend Snippet: Immunoblot analysis of CD40 in whole-cell lysates. Cell monolayers were infected with HCMV TB40E at an MOI of 1 (inf.) or mock infected (noninfected [n.i.]). Cell lysates were prepared at the indicated times and analyzed by immunoblotting. (A) CD40 protein levels were detected with a polyclonal rabbit serum directed against human CD40. (B) Blots from panel A were stripped and reprobed with a rabbit serum directed against calreticulin. (C) Signal intensities from panels A and B were densitometrically quantified; data represent the ratio of CD40 to calreticulin (y axis) versus time p.i. (x axis). (D) Protein lysates from infected or noninfected cells were treated with N-glycosidase F (+) or left untreated (−) prior to immunoblot analysis with anti-CD40 antibodies. Molecular masses are given in kilodaltons.

Article Snippet: The following monoclonal antibodies or polyclonal sera were used: mouse IgG1-fluorescein isothiocyanate (FITC)-conjugated anti-human CD54 (Calbiochem, La Jolla, Calif.), anti-human major histocompatibility complex class I (MHC-I) (BD PharMingen, San Diego, Calif.), mouse IgG1-FITC anti-human CD40 (Dianova, Hamburg, Germany), mouse IgG1-FITC anti-human CD62E (Calbiochem), antibodies G28.5 and Ro1 (anti-human CD40) (gifts from H. Engelmann, Institute for Immunology, University of Munich, Munich, Germany) ( 39 ), polyclonal rabbit anti-CD40 serum (Santa Cruz Biotechnology), rabbit antiserum against human calreticulin (Dianova), biotin-conjugated rabbit anti-mouse antibodies, and biotinylated swine anti-rabbit antibodies (Dako, Hamburg, Germany).

Techniques: Western Blot, Infection

Journal:

Article Title: AN ELISA-BASED METHOD TO QUANTIFY OSTEOCALCIN CARBOXYLATION IN MICE

doi: 10.1016/j.bbrc.2010.06.008

Figure Lengend Snippet: (A) The specificity of the different goat antibodies were tested by western blotting on dot blot membranes. GLA-OCN: fully carboxylated osteocalcin. GLU-OCN: uncarboxylated osteocalcin.

Article Snippet: GLU, GLA13 and total mouse osteocalcin ELISA Antibody Coating Buffer (CB1), ELISA Wash Buffer (WB1), General Blocker Buffer (BB1), General Assay Diluent (AD1) and Stop Solution for TMB (STOP1) were all obtained from ImmunoChemistry Technologies.

Techniques: Western Blot, Dot Blot

Journal:

Article Title: AN ELISA-BASED METHOD TO QUANTIFY OSTEOCALCIN CARBOXYLATION IN MICE

doi: 10.1016/j.bbrc.2010.06.008

Figure Lengend Snippet: (A) Quantitative measurement of GLU-OCN, GLA13-OCN and total osteocalcin in the supernatant of primary osteoblasts cultures (pOB) treated with a vehicle or with warfarin. *** p < 0.001 compared to vehicle treated osteoblasts.

Article Snippet: GLU, GLA13 and total mouse osteocalcin ELISA Antibody Coating Buffer (CB1), ELISA Wash Buffer (WB1), General Blocker Buffer (BB1), General Assay Diluent (AD1) and Stop Solution for TMB (STOP1) were all obtained from ImmunoChemistry Technologies.

Techniques:

Journal:

Article Title: AN ELISA-BASED METHOD TO QUANTIFY OSTEOCALCIN CARBOXYLATION IN MICE

doi: 10.1016/j.bbrc.2010.06.008

Figure Lengend Snippet: (A) Quantitative measurement of GLU-OCN, GLA13-OCN and total osteocalcin in serums from 2 months old wild type (WT) and Ocn −/− mice. All three forms were undetectable (UN) in Ocn −/− serums.

Article Snippet: GLU, GLA13 and total mouse osteocalcin ELISA Antibody Coating Buffer (CB1), ELISA Wash Buffer (WB1), General Blocker Buffer (BB1), General Assay Diluent (AD1) and Stop Solution for TMB (STOP1) were all obtained from ImmunoChemistry Technologies.

Techniques:

Journal: Breast Cancer Research : BCR

Article Title: Osteoblasts are “educated” by crosstalk with metastatic breast cancer cells in the bone tumor microenvironment

doi: 10.1186/s13058-019-1117-0

Figure Lengend Snippet: EOs are present in patient samples of bone metastatic breast cancer. Human patient samples of bone metastatic breast cancer were stained using multi-plex immunofluorescence for RUNX2 (green), osteocalcin (OCN, red), IL-6 (purple), and alpha-SMA (yellow). Left panel—osteoblast identification: white arrows show osteoblasts positive for both RUNX2 and OCN. Middle panel—“uneducated” and “educated” osteoblast identification: blue arrows show “uneducated” osteoblasts alpha-SMA and IL-6 positive; yellow arrows show “educated” osteoblasts alpha-SMA high, but IL-6 low; purple arrows show “educated” osteoblasts IL-6 high, but alpha-SMA low. Right panel—“educated” osteoblast identification: green arrows show “educated” osteoblasts both IL-6 and alpha-SMA low, DAPI positive. T, tumor; arrows, osteoblast. DAPI, nuclear stain. Scale bar = 50 μm

Article Snippet: The slides were incubated overnight at 4 °C with either mouse anti-human osteocalcin (R&D Systems, 10 μg/ml), rabbit anti-human RUNX2 (Abcam, 1:500), mouse anti-human alpha-smooth muscle actin (1:100, Abcam), rabbit anti-human alkaline phosphatase (1:100, Abcam), rabbit anti-human osteopontin (1:500, Abcam), or goat anti-human IL-6 (40 μg/mL, R&D Systems), Next, slides were incubated for 1 h at room temperature with either donkey anti-mouse 594, chicken anti-rabbit 594, donkey anti-goat 488, or goat anti-rabbit 488 (1:1000, Biotium).

Techniques: Staining, Immunofluorescence

Journal: Mucosal immunology

Article Title: β-Glucans in food modify colonic microflora by inducing antimicrobial protein, calprotectin, in a Dectin-1-induced-IL-17F-dependent manner.

doi: 10.1038/mi.2017.86

Figure Lengend Snippet: Figure 3 Interleukin-17F (IL-17F) production is decreased in colonic lamina propria (cLP) of Clec7a / mice. (a) Cytokine gene expression in cLP was examined by quantitative real-time reverse transcription PCR (qRT-PCR), and expression levels in Clec7a / mice are indicated relative to those in wild-type (WT) mice (n ¼ 3–8/group). (b,c) IL-17F þCD45 þ cell population in cLP layer from WT or Clec7a / mice was examined by flow cytometry. (n ¼ 4–5/group). (d) S100A8 þ and IL-17F þ cells in whole colonic cells from WT mice were examined by flow cytometry. Data are representatives of two independent experiments, and in a and c are expressed as mean±s.d. *Po0.05 vs. WT control.

Article Snippet: Antibodies against mouse CD11b (M1/70), CD11c (HL3), CD45 (30-F11), Ly6G (1A8), and Ly6C (HK1.4) were obtained from BioLegend (San Diego, CA), antibodies against IL-17F (18F10) and EpCAM-1 (G8.8) were obtained from eBioscience (San Diego, CA), and antibody against S100A8 (cat. no. BAF3059) was obtained from R&D Systems.

Techniques: Gene Expression, Reverse Transcription, Quantitative RT-PCR, Expressing, Flow Cytometry, Control

Journal: Mucosal immunology

Article Title: β-Glucans in food modify colonic microflora by inducing antimicrobial protein, calprotectin, in a Dectin-1-induced-IL-17F-dependent manner.

doi: 10.1038/mi.2017.86

Figure Lengend Snippet: Figure 6 Dectin-1 ligands are not detected in intestinal commensal microbiota. (a) Wild-type (WT) mouse fecal microbiota or Candida albicans (SC5314) were pretreated with non-immune goat or rabbit serum to block nonspecific binding. Then, they were treated with PE-labeled b-glucan- recognizing protein (Pi-BGRP) or the mutant form of the b-glucan recognition site of Pi-BGRP (Pi-BGRP.M), and PE þ population was determined by flow cytometry. (b) Thioglycollate-induced peritoneal macrophages (Mfs) from WT, Clec7a / , Myd88 / , or Clec7a / Myd88 / mice were cocultured with live Alcaligenes faecalis (multiplicity of infection (MOI): 1 or 10) for 24 h and the tumor necrosis factor-a (TNF-a) concentration in the culture supernatant was determined by enzyme-linked immunosorbent assay (ELISA; n ¼ 3/group). (c) S100A8 expression in colonic epithelial cells (cECs) from A. faecalis monocolonized (A. faecalis), germ-free (GF), or specific-pathogen-free SPF mice was examined by flow cytometry (n ¼ 3–4/group). Data in b and c are representatives of two independent experiments and are expressed as mean±s.d.

Article Snippet: Antibodies against mouse CD11b (M1/70), CD11c (HL3), CD45 (30-F11), Ly6G (1A8), and Ly6C (HK1.4) were obtained from BioLegend (San Diego, CA), antibodies against IL-17F (18F10) and EpCAM-1 (G8.8) were obtained from eBioscience (San Diego, CA), and antibody against S100A8 (cat. no. BAF3059) was obtained from R&D Systems.

Techniques: Blocking Assay, Binding Assay, Labeling, Mutagenesis, Flow Cytometry, Infection, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing

Journal: Molecules

Article Title: Gentiana lutea Extract Modulates Ceramide Synthesis in Primary and Psoriasis-Like Keratinocytes

doi: 10.3390/molecules25081832

Figure Lengend Snippet: Expression of ceramides and enzymes for the generation of long chain free fatty acids (FA) ceramides in lesional skin of psoriasis patients. ( a ) Formalin- fixed and paraffin embedded tissue of lesional skin and healthy skin was HE stained. Furthermore Ki-67, filaggrin, keratin 16 and psoriasin was stained. ( b ) Formalin- fixed and paraffin embedded tissue of lesional skin and healthy skin was stained to detect ceramides, glucosylceramides, ceramide synthase 3 (CerS3) and elongases (ELOVL 1 and ELOVL4).

Article Snippet: The following antibodies and dilutions were used for immunohistochemical stainings: polyclonal rabbit-anti-human CerS3 antibody (Antikörper-online, Aachen, Germany; 1:50), anti- ELOVL1 antibody Elabscience, Houston, TX, USA, 1:50), anti-ELOVL4 antibody (Abcam, Berlin, Germany, 1:50), anti-keratin 16 antibody (Santa Cruz, Heidelberg, Germany, 1:50), anti-Ki-67 antibody (DCS, Hamburg, Germany, 1:200), anti-filaggrin antibody (Santa Cruz, Heidelberg, Germany, 1:1000), anti-psoriasin antibody anti-ceramide antibody (Glycobiotech, Kükels, Germany, 1:25), anti-glucosylceramide antibody (Glycobiotech, Kükels, Germany, 1:50).

Techniques: Expressing, Staining