Journal: Emerging Infectious Diseases

Article Title: Livestock Susceptibility to Infection with Middle East Respiratory Syndrome Coronavirus

doi: 10.3201/eid2302.161239

Figure Lengend Snippet: Presence of MERS-CoV receptor DPP4 (IHC) and of mucosubstances (PAS) in upper and lower respiratory tract tissues from sheep, pigs, llamas, and horses. A) In the nose, DPP4 (red cytoplasmic or membrane staining) was present on the lining epithelium of pigs, llamas, and horses but not sheep. PAS staining (magenta) demonstrated more mucous cells in the lining epithelium of sheep and horses and a layer of mucus on the lining epithelium of the horses. B) DPP4 (red cytoplasmic or membrane staining) was present on the lining epithelium of the trachea, bronchus/bronchioles, and alveoli in the pigs, llamas and horses but not in the sheep. Original magnification ×400 for all images. DPP4, dipeptidyl peptidase-4; IHC, immunohistochemistry; MERS-CoV, Middle East respiratory syndrome coronavirus; PAS, periodic acid–Schiff; term., terminal.

Article Snippet: In brief, we performed DPP4 IHC staining by using 5 μg/mL of polyclonal goat IgG anti-human DPP4 antibody (R&D Systems, Abingdon, UK) and peroxidase-labeled rabbit anti-goat IgG (1:200; DAKO; Agilent Technologies Company, Santa Clara, CA, USA) as a secondary antibody.

Techniques: Membrane, Staining, Immunohistochemistry

Journal: Nature

Article Title: Dipeptidyl peptidase 4 is a functional receptor for the emerging human coronavirus-EMC.

doi: 10.1038/nature12005

Figure Lengend Snippet: Figure 4 | DPP4 is essential for virus infection. a, Inhibition of hCoV-EMC infection of Huh-7 cells by antibodies to DPP4. Supernatants collected at 2 h (open bars) and 20 h (filled bars) were tested for the presence of hCoV-EMC RNA using a TaqMan assay. Results representative of three different experiments are shown as DCt values (one-way ANOVA test, *P , 0.05; n 5 3 per group), normal goat, normal goat serum. b, Infection of human primary bronchiolar epithelial cells is blocked by the DPP4 antibodies in a dose- dependent manner and samples were analysed at 2 h (open bars) and 20 h (filled bars) after infection (one-way ANOVA test, *P , 0.05; n 5 3 per group). c, COS-7 cells transfected with plasmids encoding human DPP4 (hDPP4), bat DPP4 (bDPP4) or a control plasmid (pcDNA) were inoculated with hCoV-EMC at a multiplicity of infection of 1 and left for 1 h. Cells were washed twice and stained at 8 h after infection (original magnification, 3200) or supernatant collected at 2 h (open bars), 20 h (black bars) and 40 h (blue bars) was tested for the presence of hCoV-EMC RNA using a TaqMan assay (d). Results representative of three different experiments are expressed as GE (TCID50 ml21) values (one-way ANOVA test, *P , 0.05; n 5 4 per group). All error bars represent s.e.m.

Article Snippet: The cryosectionswere immunostained using the procedure as described26, using mouse monoclonal antibody anti-b-tubulin IV (Sigma) and goat anti-hDPP4 polyclonal antibody (R&D Systems) as primary antibodies, and Dylight-488-labelled, anti-mouse IgG (H1L) andDylight-549-labelled, anti-goat IgG (H1L) as secondary antibodies (Jackson ImmunoResearch).

Techniques: Virus, Infection, Inhibition, TaqMan Assay, Transfection, Control, Plasmid Preparation, Staining

Journal: Cell

Article Title: A bat MERS-like coronavirus circulates in pangolins and utilizes human DPP4 and host proteases for cell entry

doi: 10.1016/j.cell.2023.01.019

Figure Lengend Snippet:

Article Snippet: For the detection of hDPP4 protein in Huh-7 cells, the membrane was incubated with goat anti-hDPP4 polyclonal antibody (R&D, 0.3 μg/mL) and detected with horseradish peroxidase-conjugated rabbit anti-goat IgG (Proteintech, 1:5,000).

Techniques: Isolation, Recombinant, SYBR Green Assay, Luciferase, Reporter Assay, Sequencing, Knock-Out, Transgenic Assay, Plasmid Preparation, Software

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/JVI.00107-19

Figure Lengend Snippet: MERS-CoV S1 A binds specifically to the nasal epithelium of dromedary camels. (A) Nanoparticles displaying a multivalent MERS-CoV S1 A domain (np-S1 A ) bind to the apical surface of camel nasal ciliated epithelial cells, as revealed by red staining. np-S1 A binding is inhibited by prior neuraminidase treatment of these nasal tissues. Blank nanoparticles also do not bind to these tissues. (B) Goblet cells (arrows), visualized in purple by periodic acid-Schiff stain, are DPP4 negative, unlike nasal ciliated columnar epithelial cells (arrowheads). DPP4 expression is indicated in red. MERS-CoV S1 A binding to these goblet cells (red) can be abrogated by neuraminidase treatment. In MERS-CoV-infected camels, MERS-CoV S1 A (green) binds to both nasal ciliated columnar epithelial cells and goblet cells, while MERS-CoV N protein (red) is detected only in nasal ciliated columnar epithelial cells. np-S1 A binding is abrogated by a nanobody against the S1 A domain (Nb anti-S1 A ) but not by one against the S1 B domain (Nb anti-S1 B ). The tissues used in these experiments were sequentially cut. All pictures were taken at a ×400 magnification. (C) Nb anti-S1 A and Nb anti-S1 B bind specifically to S1 A and S1 B domains, respectively, and both bind to S1 protein, as revealed by an ELISA. The control nanobody does not bind to S1, S1 A , and S1 B . Nanobody binding is expressed as optical density at 450 nm (OD 450 ) values determined by an ELISA.

Article Snippet: MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein (Sino-Biological, Beijing, China), while DPP4 expression was detected with either 5 μg/ml goat anti-human DPP4 (R&D, Minneapolis, MN, USA) or 10 μg/ml mouse anti-human DPP4 (clone 11D7; Origene, Rockville, MD, USA).

Techniques: Staining, Binding Assay, Expressing, Infection, Enzyme-linked Immunosorbent Assay, Control

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/JVI.00107-19

Figure Lengend Snippet: Detection of MERS-CoV N protein, DPP4, α2,3-sialic acids, and MERS-CoV S1 A binding in the nasal epithelium of dromedary camels, pigs, and rabbits. MERS-CoV N protein, DPP4, α2,3-sialic acid, and MERS-CoV S1 A binding are all indicated in red. MERS-CoV N protein is detected in the nasal epithelium tissues of MERS-CoV-infected dromedary camels, pigs, and rabbits. DPP4, α2,3-sialic acid, and MERS-CoV S1 A binding were evaluated on the tissues of noninfected animals. MERS-CoV N protein and DPP4 are detected in the nasal epithelium of dromedary camel, pig, and rabbit. α2,3-Sialic acids are detected in the nasal epithelium of dromedary camel and rabbit but not in that of pig. Meanwhile, MERS-CoV S1 A binds merely to the nasal epithelium of dromedary camel. Magnification, ×400.

Article Snippet: MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein (Sino-Biological, Beijing, China), while DPP4 expression was detected with either 5 μg/ml goat anti-human DPP4 (R&D, Minneapolis, MN, USA) or 10 μg/ml mouse anti-human DPP4 (clone 11D7; Origene, Rockville, MD, USA).

Techniques: Binding Assay, Infection

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/JVI.00107-19

Figure Lengend Snippet: DPP4 expression and MERS-CoV S1 A binding in intestinal tissues of common pipistrelle bat, serotine bat, Gambian epauletted fruit bat, and Egyptian fruit bat. DPP4 expression and MERS-CoV S1 A binding are indicated in red. DPP4 is expressed at the apical surface of the intestinal epithelial cells of these four bat species. MERS-CoV S1 A binds to the apical surface of the intestinal epithelial cells of common pipistrelle bats in both villi and crypts, while in other bat species, it mostly binds to intestinal epithelial cells within the crypts. Magnification, ×400.

Article Snippet: MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein (Sino-Biological, Beijing, China), while DPP4 expression was detected with either 5 μg/ml goat anti-human DPP4 (R&D, Minneapolis, MN, USA) or 10 μg/ml mouse anti-human DPP4 (clone 11D7; Origene, Rockville, MD, USA).

Techniques: Expressing, Binding Assay

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/JVI.00107-19

Figure Lengend Snippet: MERS-CoV receptor and attachment factor in the human lower respiratory tract epithelium. (A) The MERS-CoV receptor, DPP4, is expressed in the nasal epithelium of dromedary camels, while in the lungs, it is mainly expressed in endothelial cells. In the human respiratory tract, DPP4 is expressed in bronchiolar epithelial cells (arrowhead) and type II pneumocytes (arrow) in the lungs but not in the nasal epithelium. (B) α2,3-Sialic acid expression and MERS-CoV S1 A binding are also detected in human bronchiolar epithelial cells (arrowheads) and type II pneumocytes (arrows). DPP4 expression, α2,3-sialic acids, and MERS-CoV S1 A binding are indicated in red. (C) In human alveoli, DPP4 expression (red) colocalizes in the same cells where MERS-CoV S1 A binds (green). Pictures of the nasal epithelium were taken at a ×400 magnification, and those of the alveoli were taken at a ×1,000 magnification.

Article Snippet: MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein (Sino-Biological, Beijing, China), while DPP4 expression was detected with either 5 μg/ml goat anti-human DPP4 (R&D, Minneapolis, MN, USA) or 10 μg/ml mouse anti-human DPP4 (clone 11D7; Origene, Rockville, MD, USA).

Techniques: Expressing, Binding Assay

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/JVI.00107-19

Figure Lengend Snippet: Binding of the MERS-CoV S1 A domain and MERS-CoV infection in primary normal human bronchial epithelial cells are inhibited upon prior neuraminidase treatment. (A) Removal of sialic acids using neuraminidase (NA) treatment diminishes MERS-CoV S1 A binding to primary normal human bronchial epithelial cells but not DPP4 expression in these cells. (B) The same treatment also significantly inhibits MERS-CoV infection in these cells up to 50%. Immunofluorescence images in panel A were taken at a ×400 magnification, and those in panel B were taken at a ×100 magnification. ***, P value of <0.0001 by a t test.

Article Snippet: MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein (Sino-Biological, Beijing, China), while DPP4 expression was detected with either 5 μg/ml goat anti-human DPP4 (R&D, Minneapolis, MN, USA) or 10 μg/ml mouse anti-human DPP4 (clone 11D7; Origene, Rockville, MD, USA).

Techniques: Binding Assay, Infection, Expressing, Immunofluorescence

Journal: Journal of Virology

Article Title: Species-Specific Colocalization of Middle East Respiratory Syndrome Coronavirus Attachment and Entry Receptors

doi: 10.1128/jvi.00107-19

Figure Lengend Snippet: Figure 3. DPP4 expression and MERS-CoV S1A binding in the intestinal tissues of 669

Article Snippet: 315 MERS-CoV nucleoprotein was detected with 5 μg/ml mouse anti-MERS nucleoprotein 316 (Sino-Biological, Beijing, China), while DPP4 expression with either with 5 μg/ml goat 317 anti-human DPP4 (R&D, Minneapolis, Minnesota, USA) or with 10 μg/ml mouse anti-318 human DPP4 (clone 11D7, Origene, Rockville, Maryland, USA).

Techniques: Expressing, Binding Assay