PMC:7091888 / 24514-26543 JSON TXT

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LitCovid-PubTator

LitCovid-PD-FMA-UBERON

LitCovid-PD-MONDO

LitCovid-PD-CLO

LitCovid-PD-CHEBI

LitCovid-sentences

{"project":"LitCovid-sentences","denotations":[{"id":"T143","span":{"begin":0,"end":23},"obj":"Sentence"},{"id":"T144","span":{"begin":24,"end":304},"obj":"Sentence"},{"id":"T145","span":{"begin":305,"end":620},"obj":"Sentence"},{"id":"T146","span":{"begin":621,"end":910},"obj":"Sentence"},{"id":"T147","span":{"begin":911,"end":1008},"obj":"Sentence"},{"id":"T148","span":{"begin":1009,"end":1421},"obj":"Sentence"},{"id":"T149","span":{"begin":1422,"end":1589},"obj":"Sentence"},{"id":"T150","span":{"begin":1590,"end":1812},"obj":"Sentence"},{"id":"T151","span":{"begin":1813,"end":1866},"obj":"Sentence"},{"id":"T152","span":{"begin":1867,"end":2029},"obj":"Sentence"}],"namespaces":[{"prefix":"_base","uri":"http://pubannotation.org/ontology/tao.owl#"}],"text":"Flow cytometry analysis\nFlow cytometry analysis was performed to detect the binding of SARS-CoV-2 RBD protein to hACE2 receptor in 293T cells stably expressing hACE2 (hACE2/293T).26,32 SARS-CoV and MERS-CoV RBDs, as well as 293T cells stably expressing hDPP4 receptor (hDPP4/293T), were used as controls. Briefly, cells were incubated with respective RBD of SARS-CoV-2, SARS-CoV, or MERS-CoV containing a C-terminal hFc at 20 μg/ml for 30 min at room temperature, which was followed by incubation with FITC-labeled goat anti-human IgG antibody (1:500; Thermo Fisher Scientific) for 30 min and analyzed by flow cytometry. The blockage of RBD-receptor binding was performed by incubation of soluble human ACE2 (sACE2; 5 μg/ml; R\u0026D Systems, Minneapolis, MN) receptor with respective RBD of SARS-CoV-2, SARS-CoV, or MERS-CoV (20 μg/ml), followed by the same procedure as that described above. hIgG-Fc protein (hFc: 20 μg/ml), or soluble human DPP4 (sDPP4; 5 μg/ml; R\u0026D Systems) receptor, was included as control.\nDetection of hACE2 protein expression in hACE2/293T, or hDPP4 protein expression in hDPP4/293T, stable cell lines was performed by flow cytometry analysis, as described above, except that the cells were sequentially incubated with hACE2- or hDPP4-specific goat antibody (0.5 μg/ml; R\u0026D Systems) at room temperature for 20 min and FITC-labeled anti-goat IgG antibody (1:200; Abcam, Cambridge, MA) for 1 h at 4 °C.\nFlow cytometry analysis was also performed to detect the binding between SARS-CoV-2 RBD and hACE2, or bat-ACE2 (bACE2), receptor in transiently transfected 293T cells. Briefly, 293T cells were transfected with hACE2 or bACE2 plasmid using the calcium phosphate method, and 48 h later, they were incubated with SARS-CoV-2 RBD protein at various concentrations for 30 min at room temperature. SARS-CoV and MERS-CoV RBDs were included as controls. After staining with FITC-conjugated goat anti-human IgG antibody (1:500; Thermo Fisher Scientific), the mixture was analyzed by flow cytometry as described above."}

2_test

{"project":"2_test","denotations":[{"id":"32203189-24719424-19774542","span":{"begin":179,"end":181},"obj":"24719424"},{"id":"32203189-23824801-19774543","span":{"begin":182,"end":184},"obj":"23824801"}],"text":"Flow cytometry analysis\nFlow cytometry analysis was performed to detect the binding of SARS-CoV-2 RBD protein to hACE2 receptor in 293T cells stably expressing hACE2 (hACE2/293T).26,32 SARS-CoV and MERS-CoV RBDs, as well as 293T cells stably expressing hDPP4 receptor (hDPP4/293T), were used as controls. Briefly, cells were incubated with respective RBD of SARS-CoV-2, SARS-CoV, or MERS-CoV containing a C-terminal hFc at 20 μg/ml for 30 min at room temperature, which was followed by incubation with FITC-labeled goat anti-human IgG antibody (1:500; Thermo Fisher Scientific) for 30 min and analyzed by flow cytometry. The blockage of RBD-receptor binding was performed by incubation of soluble human ACE2 (sACE2; 5 μg/ml; R\u0026D Systems, Minneapolis, MN) receptor with respective RBD of SARS-CoV-2, SARS-CoV, or MERS-CoV (20 μg/ml), followed by the same procedure as that described above. hIgG-Fc protein (hFc: 20 μg/ml), or soluble human DPP4 (sDPP4; 5 μg/ml; R\u0026D Systems) receptor, was included as control.\nDetection of hACE2 protein expression in hACE2/293T, or hDPP4 protein expression in hDPP4/293T, stable cell lines was performed by flow cytometry analysis, as described above, except that the cells were sequentially incubated with hACE2- or hDPP4-specific goat antibody (0.5 μg/ml; R\u0026D Systems) at room temperature for 20 min and FITC-labeled anti-goat IgG antibody (1:200; Abcam, Cambridge, MA) for 1 h at 4 °C.\nFlow cytometry analysis was also performed to detect the binding between SARS-CoV-2 RBD and hACE2, or bat-ACE2 (bACE2), receptor in transiently transfected 293T cells. Briefly, 293T cells were transfected with hACE2 or bACE2 plasmid using the calcium phosphate method, and 48 h later, they were incubated with SARS-CoV-2 RBD protein at various concentrations for 30 min at room temperature. SARS-CoV and MERS-CoV RBDs were included as controls. After staining with FITC-conjugated goat anti-human IgG antibody (1:500; Thermo Fisher Scientific), the mixture was analyzed by flow cytometry as described above."}