Cell lysates were prepared in lysis buffer containing 1% Nonidet P-40 and protease inhibitor cocktail (Roche) (Cao et al., 2003 (link)). Soluble proteins were immunoprecipitated using anti-Flag (M2, Sigma), anti-Myc (Sigma), or IgG of the same isotype from the same species as a negative control (Sigma). An aliquot of the total lysate (5%, vol/vol) was included as a control. Immunoblotting was performed with horseradish peroxidase (HRP)-conjugated anti-Myc (Sigma), HRP-conjugated anti-Flag (Sigma), HRP-conjugated anti-β-actin (Sigma), anti-VP35 (Creative Diagnostics), anti-IRF3 (Cell Signaling Technology), anti-phospho-IRF3 Ser396 (Cell Signaling Technology), anti-STING (Proteintech), or anti-NP (Sino Biological) antibodies. The antigen–antibody complexes were visualized via chemiluminescence (Immobilon Western Chemiluminescent HRP Substrate, Millipore). A PageRuler Western marker (Thermo) was used as a molecular weight standard.
Anti np
Manufactured by Sino Biological
The Anti-NP is a laboratory reagent used for the detection and quantification of nucleoprotein (NP) in samples. It serves as a tool for researchers and scientists to analyze the presence and levels of NP, which is a common viral protein found in various pathogens. The core function of this product is to provide a reliable and specific means of measuring NP in experimental settings.
Automatically generated - may contain errors
Lab products found in correlation
Anti vp35, by Creative Diagnostics (3 mentions)
Anti irf3, by Cell Signaling Technology (2 mentions)
Lsm 800 meta, by Zeiss (2 mentions)
Anti myc, by Merck Group (1 mentions)
Anti sting, by Proteintech (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
Anti flag m2, by Merck Group (1 mentions)
Immobilon western chemiluminescent hrp substrate, by Merck Group (1 mentions)
Hrp conjugated anti flag, by Merck Group (1 mentions)
Hrp conjugated anti β actin, by Merck Group (1 mentions)
Anti phospho irf3 ser396, by Cell Signaling Technology (1 mentions)
Hrp conjugated anti myc, by Merck Group (1 mentions)
Pageruler western marker, by Thermo Fisher Scientific (1 mentions)
Anti tbk1, by Cell Signaling Technology (1 mentions)
Anti ikkε, by Abcam (1 mentions)
Anti creb1, by Cell Signaling Technology (1 mentions)
Anti pcreb1 s133, by Abcam (1 mentions)
4 protocols using anti np
1
Immunoprecipitation and Immunoblotting Analysis
2
Immunofluorescence Imaging of Innate Immune Signaling
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Cells were transfected, fixed, permeabilized, and blocked as described above. Then, after incubation with anti-TBK1 (Cell Signaling Technology), anti-IKKε (Abcam), anti-IRF3 (Cell Signaling Technology), anti-VP35 (Creative Diagnostics), anti-NP (Sino Biological), or anti-STING (Bioss) antibodies overnight at 4°C, the cells were washed three times with phosphate buffered saline with tween 20 (PBST) buffer and then incubated with 488-conjugated anti-IRF3 (Proteintech) antibodies, FITC- or TRITC-conjugated goat anti-rabbit (or anti-mouse) IgG secondary antibodies for another 1 hr at room temperature. The cells were then stained with DAPI after washing and imaged using a laser scanning confocal microscope (Zeiss LSM 800 Meta) with a ×63 oil immersion lens.
3
SARS-CoV-2 Infection in Calu-3 Cells
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A clinical isolate nCoV-2019BetaCoV/Wuhan/WIV04/2019 was propagated in Vero E6 cells. All the infection experiments were performed in a biosafety level-3 laboratory. Briefly, Calu-3 cells were infected with SARS-CoV-2 at a multiplicity of infection of 0.05. Viral N protein and ORF3a expression in infected cells was analyzed by western blot using primary antibodies anti-NP (Sino biological, 40143-MM08) and anti-ORF3a (Bioworld Technology, NCP0017) at 48 h postinoculation.
4
Immunofluorescence Analysis of AKIP1, VP35, and CREB
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Cells were transfected, fixed, permeabilized, and blocked as described above. Then, after incubation with anti-AKIP1 (Thermo, PA5-66385, 1:100 dilution), anti-VP35 (Creative Diagnostics, CABT-B292, 1:100 dilution), anti-NP(Sino Biological, 40443-MM07 or 40443-T62, 1:100 dilution), anti-PRKACA (BD Biosciences, 610980, 1:100 dilution), anti-CREB1 (Cell Signaling Technology, 9197, 1:100 dilution), or anti-pCREB1-S133 (Abcam, ab32096, 1:50 dilution) antibodies overnight at 4 °C, the cells were washed three times with blocking solution and then incubated with FITC- or TRITC-conjugated goat anti-rabbit (or anti-mouse) IgG. The cells were then stained with DAPI after washing and imaged using a laser scanning confocal microscope (Zeiss LSM 800 Meta, built-in software ZEN2.3) with a 63× oil immersion lens. The intensity of CREB1 in the cytoplasm and nucleus was analyzed by ImageJ software.
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