Saline:Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer's Disease.
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 ◦C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer’s Disease
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 °C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Incubation:Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer's Disease.
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 ◦C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer’s Disease
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 °C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Article Title: Primary cilium and TULP3-dependent ciliary targeting of ACE2 in SARS-CoV-2 tropism
Article Snippet: .. Cells were then incubated with anti-ACE2 antibody (#AF933, R&D systems) at room temperature for 2 h, followed by incubation with anti-ARL13B antibody (#N295B/66, Developmental Studies Hybridoma Bank) overnight at 4 °C. .. Alexa anti-rabbit 488 (#A-21206, Invitrogen), Alexa anti-goat 555 (#A-21432, Invitrogen), Alexa anti-mouse 647 (#A-31571, Invitrogen) were used as secondary antibody.
Article Title: Primary cilium and TULP3-dependent ciliary targeting of ACE2 in SARS-CoV-2 tropism.
Article Snippet: .. Cells were then incubated with anti-ACE2 antibody (#AF933, R&D systems) at room temperature for 2 h, followed by incubation with anti-ARL13B antibody (#N295B/66, Developmental Studies Hybridoma Bank) overnight at 4 °C. .. Alexa anti-rabbit 488 (#A-21206, Invitrogen), Alexa anti-goat 555 (#A-21432, Invitrogen), Alexa anti-mouse 647 (#A-31571, Invitrogen) were used as secondary antibody.
Binding Assay:Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer's Disease.
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 ◦C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Article Title: Erebosis of Neurons May Exist in the Brain with Alzheimer’s Disease
Article Snippet: Coronal 20-μm sections containing the hippocampus from Bregma −2 to −4 mm were cut using a cryostat (SM2010R, Leica Biosystems, Deer Park, MD, USA) and mounted on the microscope slides. .. After being washed in Tris-buffered saline (TBS) at room temperature, sections were blocked in 10% donkey serum plus 1% bovine serum albumin in TBS containing 0.25% Triton-X 100 for 2 h at room temperature and then incubated at 4 °C overnight with the primary antibodies: anti-ACE2 antibody (1:500, rabbit, Invitrogen, catalog number: MA5-32307 or 1:500, rat, R&D System, catalog number: MAB3437, Waltham, MA, USA), anti-phospho-tau (AT8) antibody (1:500, mouse, Invitrogen, catalog number: MN1020), anti-neuronal nuclei (NeuN) antibody (1:500, mouse, Abcam, catalog number: ab104224, Cambridge, UK), anti-cleaved-caspased-3 antibody (1:500, rabbit, Cell Signaling, catalog number: 9664, Lane Danvers, MA, USA), anti-microtubule-associated protein 1A/1B light chain 3B (LC3B) antibody (1:500, rabbit, Abcam, catalog number: ab192890), anti-ionized calcium binding adaptor molecule 1 (Iba1) antibody (1:500, rabbit, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan, catalog number: 01-19741), or anti-glial fibrillary acidic protein (GFAP) antibody (1:500, goat, Abcam, catalog number: ab53554). ..
Blocking Assay:Article Title: Spike substitutions E484D, P812R and Q954H mediate ACE2-independent entry of SARS-CoV-2 across different cell lines
Article Snippet: .. For blocking of ACE2, an anti-ACE2 antibody (R&D Systems, CatAF933) was added to the cells in quadruplicate at 20 μg/ml, 10 μg/ml, 1 μg/ml, 0.1 μg/ml, and 0.01 μg/ml for 2 h. For blocking of TMPRSS2, an anti-TMPRSS2 antibody (Thermo Fisher Scientific, Cat#PA5−14264) was added to the cells in quadruplicate at 50 μg/ml, 25 μg/ml, 12.5 μg/ml, 6.25 μg/ml and 3.125 μg/ml. .. Similarly, for blocking of TMEM106B, an anti-TMEM106B antibody (Thermo Fisher Scientific, Cat#60333–1-IG) was added to the cells in quadruplicate at 50 μg/ml, 25 μg/ml, 12.5 μg/ml, 6.25 μg/ml, and 3.125 μg/ml.
Transduction:Article Title: tRNA Modifications: A Tale of Two Viruses—SARS-CoV-2 and ZIKV
Article Snippet: For SARS-CoV-2 infection of primary fibroblasts or A549 cells, the cells were previously transduced with a lentiviral vector expressing ACE2 using the lentiviral construct RRL.sin.cPPT.SFFV/Ace2.WPRE (MT136) kindly provided by Caroline Goujon (Addgene plasmid # 145842). .. Seventy-two hours after transduction, accurate ACE2 expression was controlled by Western blots using anti-ACE2 antibody (Human ACE-2 Antibody, AF933, R&D systems (a Bio-Techne brand (Minneapolis, MN, USA)). .. ZIKV virus used was either the isolate from French Polynesia (PF13), generated from molecular clone from Mathew Evans [ ], or the BeH8 (mCherry reporter) ZIKV replicon [ ] from Andres Merits (Faculty of Science and Technology, University of Tartu, Estonia).
Expressing:Article Title: tRNA Modifications: A Tale of Two Viruses—SARS-CoV-2 and ZIKV
Article Snippet: For SARS-CoV-2 infection of primary fibroblasts or A549 cells, the cells were previously transduced with a lentiviral vector expressing ACE2 using the lentiviral construct RRL.sin.cPPT.SFFV/Ace2.WPRE (MT136) kindly provided by Caroline Goujon (Addgene plasmid # 145842). .. Seventy-two hours after transduction, accurate ACE2 expression was controlled by Western blots using anti-ACE2 antibody (Human ACE-2 Antibody, AF933, R&D systems (a Bio-Techne brand (Minneapolis, MN, USA)). .. ZIKV virus used was either the isolate from French Polynesia (PF13), generated from molecular clone from Mathew Evans [ ], or the BeH8 (mCherry reporter) ZIKV replicon [ ] from Andres Merits (Faculty of Science and Technology, University of Tartu, Estonia).
Western Blot:Article Title: tRNA Modifications: A Tale of Two Viruses—SARS-CoV-2 and ZIKV
Article Snippet: For SARS-CoV-2 infection of primary fibroblasts or A549 cells, the cells were previously transduced with a lentiviral vector expressing ACE2 using the lentiviral construct RRL.sin.cPPT.SFFV/Ace2.WPRE (MT136) kindly provided by Caroline Goujon (Addgene plasmid # 145842). .. Seventy-two hours after transduction, accurate ACE2 expression was controlled by Western blots using anti-ACE2 antibody (Human ACE-2 Antibody, AF933, R&D systems (a Bio-Techne brand (Minneapolis, MN, USA)). .. ZIKV virus used was either the isolate from French Polynesia (PF13), generated from molecular clone from Mathew Evans [ ], or the BeH8 (mCherry reporter) ZIKV replicon [ ] from Andres Merits (Faculty of Science and Technology, University of Tartu, Estonia).
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