Journal: Frontiers in Immunology

Article Title: Combined p14ARF and Interferon-β Gene Transfer to the Human Melanoma Cell Line SK-MEL-147 Promotes Oncolysis and Immune Activation

doi: 10.3389/fimmu.2020.576658

Figure Lengend Snippet: Emission of immunogenic cell death markers induced by combined adenoviral p14ARF + IFNβ gene transfer. SK-MEL-147 cells transduced as previously described, incubated for 48h h before cells and supernatants were collected for ICD assays. (A) Calreticulin exposure was assessed by flow cytometry after specific antibody staining. Representative dot plots and a bar graph showing the mean and standard deviation from three independent tests with three technical replicates each. (B) Supernatant from the same cultures were collected and evaluated for ATP secretion using a luciferase-based assay (RLU, relative light units). Data represent the mean and standard deviation from at least three independent experiments. (C) Detection of secreted IFNβ protein from cell supernatant by ELISA. Data represent the mean and standard deviation from at least three independent experiments. For both (A–C) , statistical analyses were performed using one-way ANOVA test followed by the Bonferroni post-test. *p < 0.05, **p < 0.005, and ***p < 0.0005.

Article Snippet: For the evaluation of calreticulin exposure through flow cytometry, cells were probed with rabbit anti-calreticulin antibody (Novus Biologicals, Littleton, CO, USA), followed by the Alexa488-conjugated anti-rabbit secondary antibody (Thermo Fisher Scientific).

Techniques: Incubation, Flow Cytometry, Staining, Standard Deviation, Luciferase, Enzyme-linked Immunosorbent Assay

Journal: Immunology

Article Title: Acute infection of mice with Clostridium difficile leads to eIF2α phosphorylation and pro-survival signalling as part of the mucosal inflammatory response.

doi: 10.1111/imm.12122

Figure Lengend Snippet: Figure 6. Assessment of the unfolded protein response in Clostridium difficile-infected (CDI) mice. Protein lysates from the caeca and colons of untreated and C. difficile-infected mice were used to evaluate the phosphorylation of eIF2a (n = 6 pairs) (a), and the expression levels of BiP (n = 6 pairs) (d), P58IPK (n = 6 pairs) (e) and calreticulin (n = 6 pairs) (f). In each case, the panel on the left shows the image of the immunoblot for the evaluated molecule for three pairs of caeca and colons and the bar graph on the right depicts the mean SEM of the response for all the six evaluated pairs. The expression levels of Gadd34 and Wars were determined by quantitative RT-PCR (n = 12 pairs). Each bar represents the mean SEM of the relative expression of the depicted gene (b). Conventional RT-PCR was used to determine the lack of Xbp1 splicing (n = 12 pairs, of which 6 are shown) (c). A P-value ≤0.05 indicates a significant difference between the untreated and CDI samples and is marked with a ⋆.

Article Snippet: Immunoblotting antibodies against b-actin (clone 13E5), calreticulin, phospho-eIF2a (clone 119A11), eIF2a (clone L57A5), GAPDH (clone 14C10), P58IPK (clone C56E7), phospho-AKT (clone D9E), AKT (clone C67E7), phospho-STAT3 (clone D3A7) and STAT3 (clone 79D7) were obtained from Cell Signaling Technology (Danvers, MA).

Techniques: Infection, Phospho-proteomics, Expressing, Western Blot, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction

Journal: Frontiers in Pharmacology

Article Title: The pharmacological effects and safety of the raw and prepared folium of Epimedium brevicornu Maxim. on improving kidney-yang deficiency syndrome and sexual dysfunction

doi: 10.3389/fphar.2023.1233468

Figure Lengend Snippet:

Article Snippet: The eNOS antibody (ab300071), PDE5A1 antibody (ab259945), TNF-α antibody (ab1793), IL-6 antibody (ab290735), and NF-κB antibody (ab32536) were purchased from Abcam.

Techniques: CCK-8 Assay, Fluorescence, FACS, High Performance Liquid Chromatography, Concentration Assay, Saline