Journal: mBio

Article Title: The Vi Capsular Polysaccharide of Salmonella Typhi Promotes Macrophage Phagocytosis by Binding the Human C-Type Lectin DC-SIGN

doi: 10.1128/mbio.02733-22

Figure Lengend Snippet: Vi capsulated S. Typhi binds human DC-SIGN. (A and G) Wells were coated with formalin-killed capsulated S. Typhi (wild type [wt]) or formalin-killed noncapsulated S. Typhi ( tviB-vexE ), and binding of the indicated murine (m) or human (h) CLR proteins fused to the Fc portion of human IgG 1 (hFc) was assessed by ELISA. Graphs show signal (OD 450 ) generated by binding to wt-coated wells which was higher than the background levels generated by binding to tviB-vexE -coated wells. (A) Bars indicate geometric means ± standard error from 4 biological repeats. (B) Binding of hFc or hDC-SIGN-hFc fusion protein (red fluorescence) to the surface of GFP-labeled capsulated S. Typhi (green fluorescence) was visualized by fluorescence microscopy. (C) Binding of hFc or hDC-SIGN-hFc fusion protein (green fluorescence) to the surface of S. Typhi (Hoechst DNA stain, blue fluorescence) synthesizing the Vi capsular antigen, detected by staining with anti-Vi antibodies (red fluorescence), was visualized by fluorescence microscopy. (D and E) Human cervical carcinoma epithelial (HeLa) cells were either mock treated (mock) or transfected with a plasmid encoding CD209 . (D) Transcription levels of CD209 were measured using quantitative real-time PCR. (E) The number of capsulated S. Typhi (CFU) adhering to HeLa cells was determined. (D and E) Bars indicate geometric means ± standard error from 3 biological repeats. (F) Streptavidin-coated microplates were coated with Biotin-Vi or biotin, and binding to hDC-SIGN-hFc was assessed by ELISA in the presence (+) or absence (–) of EDTA. Graphs show signal (OD 450 ) generated by binding to biotin-Vi-coated wells that was higher than background levels generated by binding to biotin-coated wells. Bars indicate geometric means ± standard error from 5 biological repeats. (G) Binding to hDC-SIGN-hFc was assessed by ELISA in the presence (+) or absence (–) of EDTA. Bars indicate geometric means ± standard error from 3 biological repeats. *, P < 0.05; **, P < 0.01.

Article Snippet: To express DC-SIGN in HeLa cells, an untagged DC-SIGN expression vector (HG10200-UT, Sino Biological) was transfected.

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Generated, Fluorescence, Labeling, Microscopy, Staining, Transfection, Plasmid Preparation, Real-time Polymerase Chain Reaction

Journal: Frontiers in Immunology

Article Title: Human surfactant protein D facilitates SARS-CoV-2 pseudotype binding and entry in DC-SIGN expressing cells, and downregulates spike protein induced inflammation

doi: 10.3389/fimmu.2022.960733

Figure Lengend Snippet: Binding of SARS-CoV-2 Spike-Pseudotypes to DC-SIGN expressing cells. DC-HEK cells were incubated with SARS-CoV-2 Spike Pseudotypes for 30 min at 37°C. Spike Pseudotypes challenged DC-HEK cells were fixed with 4% paraformaldehyde, washed, and blocked with 5% FCS. The cells were probed with rabbit anti-SARS-CoV-2 Spike antibody and mouse anti-DC-SIGN to detect the presence of Spike-Pseudotypes and DC-SIGN expressed on the cells, respectively. Alexa Fluor 647 conjugated goat anti-mouse antibody (Abcam), Alexa fluor 488 conjugated goat anti-rabbit antibody (Abcam), and Hoechst (Invitrogen, Life Technologies) were used to detect the primary antibodies and nucleus.

Article Snippet: HEK 293T cells were transiently transfected with a plasmid expressing human DC-SIGN (HG10200-UT; Sino Biological), using Promega FuGENE™ HD Transfection Reagent (Fisher Scientific).

Techniques: Binding Assay, Expressing, Incubation

Journal: Frontiers in Immunology

Article Title: Human surfactant protein D facilitates SARS-CoV-2 pseudotype binding and entry in DC-SIGN expressing cells, and downregulates spike protein induced inflammation

doi: 10.3389/fimmu.2022.960733

Figure Lengend Snippet: rfhSP-D promotes interaction between SARS-CoV-2 Spike Pseudotypes and DC-SIGN expressing cells. DC-HEK cells (A) and DC-THP-1 cells (B) were treated with rfhSP-D and SARS-CoV-2 Spike-Pseudotypes. The cell binding was analysed using Alexa Fluor 488 (FTIC) and Alexa Fluor 647 (APC); the fluorescence intensity was measured using a GloMax 96 Microplate Luminometer (Promega). An increased fluorescence intensity was observed in DC-HEK and DC-THP-1 cells treated with 20 µg/ml of rfhSP-D compared to cells challenged with Spike pseudotypes alone. Experiments were conducted in triplicates, and error bars represent ± SEM. Unpaired t-test was used to calculate the significance (*p < 0.05, **p < 0.01, and ***p < 0.001) (n = 3). (0, untreated sample; 20, treated sample).

Article Snippet: HEK 293T cells were transiently transfected with a plasmid expressing human DC-SIGN (HG10200-UT; Sino Biological), using Promega FuGENE™ HD Transfection Reagent (Fisher Scientific).

Techniques: Expressing, Binding Assay, Fluorescence

Journal: Cell reports

Article Title: TRIM5α Restricts Flavivirus Replication by Targeting the Viral Protease for Proteasomal Degradation

doi: 10.1016/j.celrep.2019.05.040

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: DC-SIGN expression plasmid , Sino Biological , HG10200-UT.

Techniques: Transduction, Recombinant, Modification, Protease Inhibitor, Staining, Transfection, Clone Assay, Construct, shRNA, Sequencing, Luciferase, Plasmid Preparation, Expressing, Software, Imaging, Western Blot