goat anti human ace2 ectodomain antibody  (R&D Systems)

Journal: The Journal of general virology

Article Title: Replication-dependent downregulation of cellular angiotensin-converting enzyme 2 protein expression by human coronavirus NL63.

doi: 10.1099/vir.0.043919-0

Figure Lengend Snippet: Fig. 1. Downregulation of ACE2 expression by HCoV-NL63. (a) Chemiluminescence detection of ACE2 protein expression in mock- or HCoV-NL63-infected LLC-MK2 cells cultured at 34 6C. (b) b-Actin protein expression in mock- or HCoV-NL63- infected cultures as loading control. (c) The HCoV-NL63 viral yield measured in the culture supernatant. (d) Confocal images of DNA (blue), ACE2 (green) and HCoV-NL63 N protein (red) in mock- and HCoV-NL63-infected LLC-MK2 cells 5 days p.i. The results from a representative experiment are shown.

Article Snippet: Coverslips were stained with serum from HCoV-NL63 nucleocapsid (N) protein immunized rabbits (1 : 400; Eurogentec) and goat polyclonal anti-human ACE2 ectodomain (2 mg ml21, AF993; R&D Systems), followed by incubation with donkeyderived, anti-rabbit IgG and anti-goat IgG [(H+L), 7.5 mg ml21; Jackson Immunoresearch], conjugated with Dylight 649 and Dylight 594, respectively.

Techniques: Expressing, Infection, Cell Culture, Control

Journal: The Journal of general virology

Article Title: Replication-dependent downregulation of cellular angiotensin-converting enzyme 2 protein expression by human coronavirus NL63.

doi: 10.1099/vir.0.043919-0

Figure Lengend Snippet: Fig. 2. ACE2 down modulation is dependent on HCoV-NL63 replication efficiency. (a) The HCoV-NL63 viral yield monitored at 34 and 37 6C. (b) The ACE2 concentration at 34 6C in mock- and HCoV-NL63-infected cultures (**5P,0.01, ****5P,0.0001). (c) The ACE2 concentration at 37 6C in mock- and HCoV-NL63-infected cultures. (d) Human metapneumovirus (hMPV) yield monitored at 34 6C. (e) The percentage of ACE2 expression in mock- and hMPV-infected cultures (f) ACE concentration at 34 and 37 6C in mock- and HCoV-NL63-infected cultures. The results are shown as the mean of two independent experiments with SEM, except for panel (d) and (e) where the results from a representative experiment are shown.

Article Snippet: Coverslips were stained with serum from HCoV-NL63 nucleocapsid (N) protein immunized rabbits (1 : 400; Eurogentec) and goat polyclonal anti-human ACE2 ectodomain (2 mg ml21, AF993; R&D Systems), followed by incubation with donkeyderived, anti-rabbit IgG and anti-goat IgG [(H+L), 7.5 mg ml21; Jackson Immunoresearch], conjugated with Dylight 649 and Dylight 594, respectively.

Techniques: Concentration Assay, Infection, Expressing

Journal: Virology

Article Title: A study on antigenicity and receptor-binding ability of fragment 450–650 of the spike protein of SARS coronavirus

doi: 10.1016/j.virol.2006.09.022

Figure Lengend Snippet: Co-precipitation of recombinant S450–650 with ACE2 in the lysate of Vero E6 cells. Vero E6 cells (1 × 10 6 ) were sequentially treated with goat anti-human ACE2 Abs, or goat IgG (2 μg), and FITC-conjugated rabbit anti-goat IgG. The stained cells were subjected to flow cytometric analysis for determination of ACE2 expression on the cell surface (A). For the co-precipitation experiment (B), Vero E6 cells (2.5 × 10 7 ) were treated with, or without, recombinant S450–650 (15 μg/ml) at 4 °C for 1 h. After washes, the cells were lysed using lysis buffer and the lysate treated with PT31 serum followed by protein-A/G-agarose precipitation. The precipitated materials were run on SDS-PAGE 12% gels and the protein bands transferred to nitrocellulose membranes for WB assays using goat anti-human ACE2 Abs as the first Abs, the detection Abs were HRP-labeled rabbit anti-goat IgG. In a parallel experiment (C), lysate of the S450–650-treated, or untreated, Vero E6 cells were precipitated using goat anti-human ACE2 Abs and protein-A/G-agarose. The precipitates were assayed by WB using PT31 serum (first Ab) and HRP-labeled goat anti-human IgG (detection Ab).

Article Snippet: Goat anti-human ACE2 ectodomain polyclonal Abs was purchased from R&D systems (Minneapolis, MN, USA).

Techniques: Recombinant, Staining, Expressing, Lysis, SDS Page, Labeling

Journal: Virology

Article Title: A study on antigenicity and receptor-binding ability of fragment 450–650 of the spike protein of SARS coronavirus

doi: 10.1016/j.virol.2006.09.022

Figure Lengend Snippet: ACE2 interaction with the β5 and β6 strands of S-RBM. (A) Crystal structure of the RBD in complex of human receptor ACE2 as reported by Li and co-workers . Details of the binding interface between S-RBM and the ACE2 molecule and calculation results of the contact area and binding energy are shown in panels B and C, respectively. The S protein-binding areas of the ACE2 molecule (purple) are colored in green and the β5 and β6 strands in gold. The binding energy here is the energy required to disassemble the RBD-ACE2 complex into separate parts. The figure is prepared using PYMOL and contact area and binding energy calculated by SPDBV. PDB file accession number is 2AJF.

Article Snippet: Goat anti-human ACE2 ectodomain polyclonal Abs was purchased from R&D systems (Minneapolis, MN, USA).

Techniques: Binding Assay, Protein Binding

Journal: Microbes and Infection

Article Title: Establishment of Vero E6 cell clones persistently infected with severe acute respiratory syndrome coronavirus

doi: 10.1016/j.micinf.2005.05.013

Figure Lengend Snippet: ACE2 down-regulation on the surface of Vero/SARS-CoV cells. Mock-infected Vero E6 cells and persistently infected Vero/SARS-CoV that had been cultured for 5 months, as well as cell clones #21, #12 and #13 that had also been cultured for 3 months after their preparation were stained without fixation, with anti-ACE2 PAb as well as anti-SARS-CoV S murine MAb (3A2). The stained cells were subjected to IF microscopy and flow cytometry.

Article Snippet: For detection of cell surface ACE2, goat ACE2 PAb (anti-human ACE2 ectodomain antibody; Research and Diagnostic Systems, Inc.) was used.

Techniques: Infection, Cell Culture, Clone Assay, Staining, Microscopy, Flow Cytometry