The following reagents were purchased from the indicated sources: anti-α-tubulin antibody (clone DM1A), 12-O-tetradecanoyl-13-acetate (TPA), paclitaxel, and nocodazole (Sigma-Aldrich, St. Louis, MO, USA); 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI; Nacalai Tesque, Kyoto, Japan); Dynabeads® Protein G, Alexa Fluor™ Plus 647-conjugated anti-mouse IgG antibody, Alexa Fluor™ Plus 555-conjugated anti-mouse IgG antibody, Alexa Fluor™ 488-conjugated phalloidin, and pHrodo™ Green Zymosan A BioParticles™ Conjugate (Invitrogen, Oslo, Norway); AcidiFluor™ ORANGE-NHS (AFO: Goryo Chemical, Hokkaido, Japan); formaldehyde (Polysciences, Warrington, PA, USA).
4 6 diamidino 2 phenylindole dihydrochloride dapi
Manufactured by Nacalai Tesque
Sourced in Japan
4',6-diamidino-2-phenylindole dihydrochloride (DAPI) is a fluorescent dye that binds to DNA. It is commonly used for staining and visualizing cell nuclei in fluorescence microscopy.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 700, by Zeiss (4 mentions)
Goat igg bs 0294p, by Bioss Antibodies (2 mentions)
Rabbit igg bs 0295p, by Bioss Antibodies (2 mentions)
Nocodazole, by Merck Group (1 mentions)
Alexa fluor 488 conjugated phalloidin, by Thermo Fisher Scientific (1 mentions)
Anti α tubulin antibody clone dm1a, by Merck Group (1 mentions)
Paclitaxel, by Merck Group (1 mentions)
Formaldehyde, by Polysciences (1 mentions)
Bz 8000, by Keyence (1 mentions)
Ab53554, by Abcam (1 mentions)
Ab49873, by Abcam (1 mentions)
Ab150129, by Abcam (1 mentions)
Blocking one histo, by Nacalai Tesque (1 mentions)
Ab150074, by Abcam (1 mentions)
Aqua poly mount, by Polysciences (1 mentions)
Foxa2, by Cell Signaling Technology (1 mentions)
Sox17, by R&D Systems (1 mentions)
Alexa 488 conjugated anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Sars cov 2 nucleocapsid, by GeneTex (1 mentions)
Sars spike glycoprotein, by Abcam (1 mentions)
5 protocols using 4 6 diamidino 2 phenylindole dihydrochloride dapi
1
Immunofluorescence Staining Protocol
2
Influenza virus infection in MDCK cells
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MDCK cells (7.6 × 105 cells) in glass base dishes (IWAKI, Asahi Glass Co. Ltd., Chiba, Japan) were infected with MFPTr virus or the WT virus at 1 PFU/cell or mock infected with PBS. At 0, 5, 10, and 15 hours post-infection, the infected cells were fixed using 4% paraformaldehyde phosphate buffer solution, and permeabilized with 0.2% Triton X-100. After blocking treatment with 1% BSA in PBS, the cells were immune-reacted with FITC conjugated-mouse anti-influenza A virus nucleoprotein (431) antibody (Abcam PLC, Cambridge, UK), and nuclei were counterstained by 4’,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Nacalai Tesque, Kyoto, Japan). Stained cells were visualized using fluorescence microscopy, BZ-8000 (Keyence Corp., Osaka, Japan), and obtained images were analyzed by using ImageJ [12 (link)].
3
Immunohistochemical Analysis of TG and spinal trigeminal nucleus caudalis
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Immunohistochemical staining of TG and spinal trigeminal nucleus caudalis was done following our previously published work [13 (link)]. Briefly, the sections from TG were stained with primary antibody cleav Casp3 and secondary antibody (Table 3 ). The spinal trigeminal nucleus caudalis sections were stained with primary antibodies C-Fos and GFAP followed by secondary antibodies (Table 3 ). Nuclear staining was done using 4′,6-Diamidino-2-phenylindole dihydrochloride (DAPI, 1:100, Nacalai Tesque, Inc., Kyoto, Japan). Isotypes (goat IgG bs-0294P, and rabbit IgG bs-0295P, Bioss Antibodies, Boston, MA, USA) and only secondary antibodies were used as positive and negative control. Images were observed and acquired by confocal laser-scanning microscope (LSM 700, Carl Zeiss, Oberkochen, Germany).
4
Trigeminal Ganglion Immunostaining Protocol
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Six to 8 microns-thick frozen sections of trigeminal ganglion were fixed in 4% paraformaldehyde, permeabilized with 0.1% Triton and subsequently blocked with Blocking One Histo (Nacalai Tesque, Kyoto, Japan). Sections were incubated with primary antibodies, rabbit anti-glutamine synthetase (1:1000; ab49873, Abcam, Cambridge, UK) and goat anti-GFAP (1:500; ab53554, Abcam), overnight at 4 °C. Sections were incubated with secondary antibodies for 2 h at room temperature using donkey anti-goat IgG Alexa Fluor 488 (1:200; ab150129, Abcam) and donkey anti-rabbit Alexa Fluor 555 (1:200; ab 150074, Abcam). For nuclear staining, 4’, 6-Diamidino-2-phenylindole dihydrochloride (DAPI, 1:100, Nacalai Tesque, Inc., Kyoto, Japan) was used for 15 min at room temperature, followed by mounting with Aqua-Poly/Mount (Polysciences, Inc., Warrington, PA, USA). Isotypes (goat IgG bs-0294P, and rabbit IgG bs-0295P, Bioss Antibodies, Boston, MA, USA) and only secondary antibodies were used as positive and negative control. Images were observed and acquired by confocal laser-scanning microscope (LSM 700, Carl Zeiss, Oberkochen, Germany).
5
Comprehensive Immunocytochemical Staining Assay
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Cell staining was performed as previously described (Hirata et al., 2014) . Briefly, cells were fixed, permeabilized, blocked, and incubated with primary antibodies against SOX17 (1:500; R&D Systems), FOXA2 (1:1000; Cell Signaling Technology), CPM (1:400; Fujifilm Wako), TTF1/NKX2-1 (1:400; Abcam, Cambridge, UK), mature-SFTPB (1:500; Seven Hills Bioreagents, Cincinnati, OH, USA), pro-SFTPC (1:200; Abcam), SCGB3A2 (1:500; Abcam), and ACE2 (1:100; R&D systems), SARS-CoV-2 nucleocapsid (1:100; GeneTex, Irvine, CA, USA), or SARS spike glycoprotein (1:100; Abcam) in 5% FBS in PBS overnight at 4°C. After washing with PBS, cells were incubated with secondary antibodies, Alexa 488-conjugated anti-mouse IgG, Alexa 488-cojugated anti-goat IgG, and Alexa 594 -conjugated anti-rabbit IgG (1:200; Thermo Fisher Scientific) in 5% FBS for 1 hr at room temperature. Nuclei were counterstained with 4,6-Diamidino-2-phenylindole dihydrochloride (DAPI; Nacalai Tesque, Kyoto, Japan). Cells were mounted with VEC-TACHIELD mounting medium (Vector Laboratories, Burlingame, CA, USA). Fluorescence images were captured using a BIOREVO BZ-9000 fluorescence microscope (Keyence, Osaka, Japan) or a Nikon A1 confocal microscope (Nikon, Tokyo, Japan).
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