All compounds except for ciclesonide and EIDD-1931 were purchased from MedChemExpress (Monmouth Junction, NJ, USA). ciclesonide and EIDD-1931 were purchased from Cayman Chemical (Ann Arbor, MI, USA). The following are the lot numbers and purities of each compound: aprotinin (lot, 62009; purity of ≥98.0%), bromhexine hydrochloride (lot, 15159; purity of 99.39%), niclosamide (lot, 15718; purity of 98.68%), ciclesonide (lot, 0472344-2; purity of ≥98.0%), remdesivir (lot, 46182; purity of 99.78%), EIDD-2801 (lot, 67548; purity of 99.94%), and EIDD-1931 (lot, 0590872-1; purity of ≥95.0%). Stock solution was dissolved in dimethyl sulfoxide (DMSO) at a 10 mM concentration. Anti‐SARS‐CoV‐2 N protein antibody was purchased from Sino Biological Inc. (Beijing, China). Alexa Fluor 488 goat anti‐rabbit IgG (H + L) secondary antibody and Hoechst 33342 were purchased from Molecular Probes. Paraformaldehyde (PFA) (32% aqueous solution) and normal goat serum were purchased from Electron Microscopy Sciences (Hatfield, PA, USA) and Vector Laboratories, Inc. (Burlingame, CA, USA), respectively.
Alexa fluor 488 goat anti rabbit igg h l secondary antibody
Manufactured by Thermo Fisher Scientific
Sourced in United States, China
Alexa Fluor 488 goat anti-rabbit IgG (H + L) secondary antibody is a fluorescently-labeled secondary antibody that binds to rabbit primary antibodies. It is designed for detection and visualization applications in immunoassays and other analytical techniques.
Automatically generated - may contain errors
Lab products found in correlation
Hoechst 33342, by Thermo Fisher Scientific (6 mentions)
Anti sars cov 2 n protein antibody, by Sino Biological (5 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (3 mentions)
Eidd 1931, by MedChemExpress (2 mentions)
Ciclesonide, by Cayman Chemical (2 mentions)
Eidd 1931, by Cayman Chemical (2 mentions)
Paraformaldehyde pfa 32 aqueous solution, by Electron Microscopy Sciences (2 mentions)
Normal goat serum (ngs), by Electron Microscopy Sciences (2 mentions)
Ciclesonide, by MedChemExpress (2 mentions)
Confocal microscope, by Leica (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (2 mentions)
Normal goat serum (ngs), by Vector Laboratories (2 mentions)
Chloroquine, by Merck Group (2 mentions)
Chloroquine, by Welgene (2 mentions)
Dulbecco s phosphate buffered saline dpbs, by Welgene (2 mentions)
Anti rabbit igg h l hrp conjugated antibody, by Promega (2 mentions)
Hrp conjugated rabbit anti 6 his antibody a190 114p, by Fortis Life Sciences (2 mentions)
Lsm 700, by Zeiss (2 mentions)
Axio observer z1, by Zeiss (2 mentions)
Blocking reagent, by Roche (2 mentions)
20 protocols using alexa fluor 488 goat anti rabbit igg h l secondary antibody
1
SARS-CoV-2 Anti-Viral Compound Screening
2
Immunofluorescent Labeling of Microglia
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Cryosections of the brain were first fixed in 4% PFA for 15 min, then incubated in 0.2% Triton X-100 in PBS for 10 minutes. Blocking the sections with 10% goat serum for an hour at RT and then Iba-1 antibody (1:500 dilution, Cell Signaling Technology, MA, USA) was incubated on the sections overnight at 4°C after three PBS washes. Following three PBS washes, the sections were incubated for 1 hour at RT with Alexa Fluor® 488 goat anti-rabbit IgG (H + L) Secondary antibody (1:500 dilution, Molecular Probes, Eugene, Oregon, United States) in PBS containing 1% goat serum. The sections underwent three PBS washes before being counterstained with DAPI (1:1,000 dilution, Beyotime Institute of Biotechnology) in PBS for 5 min at RT. A Leica confocal microscope (Leica Microsystems, Wetzlar, Germany) was used to capture the fluorescence pictures, which were then processed by ImageJ.
3
Anti-SARS-CoV-2 N Protein Antibody Localization
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Plitidepsin (batch#: 16 D19) and ampoule (batch#: 60108) were provided by Boryung Pharmaceutical. Anti-SARS-CoV-2 N protein antibody was purchased from Sino Biological lnc. Alexa Fluor 488–goat anti-rabbit [IgG (H+L) secondary antibody and Hoechst 33342 were purchased from Molecular Probes.
4
Anti-SARS-CoV-2 N Protein Assay
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Isoginkgetin (MW-566.51 g/mol), % purity ≥ 98% (HPLC) and afzelin (432.38 g/mol), % purity ≥ 90% (LC/MS-UV) were purchased from Sigma-Aldrich, and chloroquine, lopinavir, and remdesivir (used as reference compounds) were bought from Sigma-Aldrich, SelleckChem, and MedChemExpress, respectively. The anti-SARS-CoV-2 N protein antibody was purchased from Sino Biological Inc. (China), with Hoechst-33342 and the Alexa Fluor-488 goat anti-rabbit IgG (H + L) secondary antibody purchased from Molecular Probes. Also, paraformaldehyde (PFA) (a 32% aqueous solution) and normal goat serum were obtained from Vector Laboratories, Inc. (Burlingame, CA) and Electron Microscopy Sciences (Hatfield, PA), respectively. chloroquine was mixed in Dulbecco's Phosphate-Buffered Saline (DPBS; Welgene). For in vitro studies, all reagents were well mixed in DMSO.
5
SARS-CoV-2 Anti-Viral Compound Screening
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All compounds except for ciclesonide and EIDD-1931 were purchased from MedChemExpress (Monmouth Junction, NJ, USA). ciclesonide and EIDD-1931 were purchased from Cayman Chemical (Ann Arbor, MI, USA). The following are the lot numbers and purities of each compound: aprotinin (lot, 62009; purity of ≥98.0%), bromhexine hydrochloride (lot, 15159; purity of 99.39%), niclosamide (lot, 15718; purity of 98.68%), ciclesonide (lot, 0472344-2; purity of ≥98.0%), remdesivir (lot, 46182; purity of 99.78%), EIDD-2801 (lot, 67548; purity of 99.94%), and EIDD-1931 (lot, 0590872-1; purity of ≥95.0%). Stock solution was dissolved in dimethyl sulfoxide (DMSO) at a 10 mM concentration. Anti‐SARS‐CoV‐2 N protein antibody was purchased from Sino Biological Inc. (Beijing, China). Alexa Fluor 488 goat anti‐rabbit IgG (H + L) secondary antibody and Hoechst 33342 were purchased from Molecular Probes. Paraformaldehyde (PFA) (32% aqueous solution) and normal goat serum were purchased from Electron Microscopy Sciences (Hatfield, PA, USA) and Vector Laboratories, Inc. (Burlingame, CA, USA), respectively.
6
Anti-SARS-CoV-2 Protein Detection Assay
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Isolated CBDs were used, and lopinavir (LPV; S1380) was purchased from SelleckChem and remdesivir (HY- 104077) from MedChemExpress, and chloroquine was obtained from Sigma-Aldrich. chloroquine was mixed in Dulbecco's Phosphate-Buffered Saline (DPBS; Welgene). For in vitro studies, all reagents were well mixed in DMSO. The Anti-SARS-CoV-2 N protein antibody was obtained from Sino Biological Inc. (China), while Hoechst-33342 and Alexa Fluor-488 goat anti-rabbit IgG (H + L) secondary antibody were bought from the Molecular Probes. Also, paraformaldehyde (PFA) (32% aqueous solution) and normal goat serum were purchased from Vector Laboratories, Inc. (Burlingame, CA) and Electron Microscopy Sciences (Hatfield, PA), respectively.
7
Immunofluorescence Staining of Brain Cryosections
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Brain cryosections were fixed in 4% PFA for 15 min, followed by incubation with 0.2% Triton X-100 in PBS for 10 min. After washes with PBS three times, the sections were blocked with 10% goat serum for 1 h at room temperature and then incubated with Iba-1 antibody (1:500 dilution, Wako, Osaka, Japan) or COX2 antibody (1:200 dilution, Abcam) in PBS containing 1% goat serum overnight at 4°C. After three washes with PBS, the sections were incubated with Alexa Fluor® 488 goat anti-rabbit IgG (H + L) Secondary antibody (1:500 dilution, Molecular Probes, Eugene, Oregon, United States) in PBS containing 1% goat serum for 1 h at room temperature. After three washes with PBS, the sections were counterstained with DAPI (1:1,000 dilution, Beyotime Institute of Biotechnology) in PBS for 5 min at room temperature. The fluorescence images were photographed using a Leica confocal microscope (Leica Microsystems, Wetzlar, Germany), which were analyzed by ImageJ.
8
SARS-CoV-2 Antibody Detection Protocol
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Antibodies to the nucleocapsid (N) protein and spike (S) domains were obtained from Thermo Fisher Scientific (Waltham, MA, USA) and Sino Biological (Wayne, PA, USA). IRF3 (#4962), phospho-IRF3 (#4947S), and STAT1 (#9172) were purchased from Cell Signaling Technology Inc. (Danvers, MA, USA), GAPDH (sc-25778) and phosphor-STAT1 (sc-7988) were purchased from Santa Cruz Biotechnology Inc. (Dallas, TX, USA). Alexa Fluor 488 goat anti-rabbit IgG (H + L) secondary antibody was purchased from Invitrogen (Carlsbad, CA, USA). Vectashield mounting medium with DPAI (H-1500) was purchased from Vector Labs (Burlingame, CA, USA).
9
Antibody Panel for Chicken Immune Signaling
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Mouse monoclonal GFPuv/eGFP antibody was purchased from R&D Systems (Minneapolis, MN, USA). The following reagents were from the indicated manufacturers: mouse anti-chicken MHC Class I-PE and mouse anti-chicken β2m-PE antibody (Southern Biotech, Birmingham, AL, USA); rabbit anti-chicken STAT1 (phospho-Ser727), anti-chicken STAT3 (phospho-Ser727), and anti-chicken JAK2 (phospho-Tyr1007/Tyr1008) antibody (Santa Cruz Biotech Dallas, Texas, USA); rabbit anti-chicken SOCS1, anti-chicken STAT1, anti-chicken STAT3 antibodies, horseradish peroxidase (HRP )-linked anti-rabbit secondary antibodies, and protein G–sepharose bead (Sigma-Aldrich, Louis, MO, USA); rabbit anti-chicken SHP2 (phospho-Tyr542), anti-chicken JAK2, and anti-chicken TYK2 antibody (Biorbyt, San Francisco, CA, USA); rabbit anti-chicken GAPDH antibody (Abcam, Cambridge, MA, USA); Alexa Fluor® 488 Goat anti-rabbit IgG (H+L) secondary antibody (Invitrogen, Carlsbad, CA, USA); mouse monoclonal anti-chicken IFN-γ, IL-17A, IL-12p40 antibody and recombinants of these proteins (kindly provided by Dr. Hyun S. Lillehoj, USDA). EZ-LinkTM Sulfo-NHS-LC-Biotin, goat anti-mouse IgG secondary antibody linked HRP conjugate, and HRP-conjugated streptavidin were purchased from Thermo Scientific (Waltham, MA, USA).
10
Phosphorylation-Specific Antibody Assay
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TAK1 (phosphor-Ser192) (#orb7051) and NFKB1 (phosphor-Ser933) antibodies were purchased from Biorbyt (Cambridge, UK). The Anti-NF-kappaB p105 (pS932) phospho antibody (#MBS8210747) was purchased from MyBioSource (San Diego, CA). Mouse anti-chicken glyceraldehyde-3-phosphate dehydrogenase (GAPDH ) antibody (#AM4300), goat anti-mouse IgG horseradish peroxidase (HRP )-conjugated antibody (#A16078), and radioimmunoprecipitation assay (RIPA ) lysis and extraction buffers were purchased from Thermo Fisher Scientific (Waltham, MA). Alexa Fluor 488 goat anti-rabbit IgG (H+L) secondary antibody and 4,6-diamidino-2-phenylindole (DAPI ) were purchased from Invitrogen (#A-11008; Carlsbad, CA). Anti-rabbit IgG (H+L) HRP-conjugated antibody was purchased from Promega (#W4011; Madison, WI). The CD9 antibody was purchased from Cell Signaling (#13174; Danvers, MA).
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