For intracellular distribution of CD63 in the hBMSCs after incubation for 48 h on different titanium topographies, cells were fixed with 4% paraformaldehyde for 10 min and blocked with PBS containing 0.1% Triton X-100 and 1% bovine serum albumin for 30 min. The cells on the titanium samples were first incubated with rabbit anti-CD63 (1:200; Immunoway) at 4 °C overnight and stained with goat anti-rabbit Alexa Fluor 488 IgG H&L antibody (1:500; Abcam) and DAPI-containing mounting medium (Beyotime). Images of the immunofluorescent staining were captured with a confocal laser scanning microscope (LSM780, Carl Zeiss Meditec AG, Germany).
Dapi containing mounting medium
Manufactured by Beyotime
Sourced in China
DAPI-containing mounting medium is a laboratory reagent used to mount and preserve biological samples for microscopy analysis. It serves the core function of immobilizing and protecting samples while staining nucleic acids with the fluorescent dye DAPI (4',6-diamidino-2-phenylindole) for visualization.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 780, by Zeiss (2 mentions)
Spinsr10, by Olympus (1 mentions)
Alexa fluor 488 goat anti, by Thermo Fisher Scientific (1 mentions)
Dylight 488 labeled secondary antibody, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 594 conjugated secondary ab, by Thermo Fisher Scientific (1 mentions)
Lsm 800, by Zeiss (1 mentions)
Alexa fluor 594 conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
Confocal laser scanning microscope, by Leica (1 mentions)
Lsm510 meta confocal laser scanning microscope, by Leica (1 mentions)
4 protocols using dapi containing mounting medium
1
Intracellular CD63 Distribution in hBMSCs on Titanium
2
Immunostaining Procedure for NLRP3, KAT5, GOLGA4, TGN38
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For immunostaining, cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100 in PBS, then the cells were incubated with primary antibodies(anti-NLRP3 antibody (1:100), anti-KAT5 antibody (1:100), anti-GOLGA4(1:200) or anti-TGN38(1:200)) followed by staining with secondary antibody(DyLight 488-labeled secondary antibody (Invitrogen)(1:50),Alexa Fluor 594-conjugated secondary Ab (Invitrogen) (1:50),Alexa Fluor® 488 Goat anti-mouse IgG (minimal x-reactivity) Antibody(1:100) or Cy3–conjugated Affinipure Goat Anti-Rabbit IgG(H + L) (1:100)), while nuclei were stained with DAPI containing mounting medium (Beyotime). To better preserve the dTGN structures in COS-7 cells, 0.01% Triton X-100 and 0.1% saponin were respectively in place of 0.1% Triton X-100 in permeabilization step for immunostaining of GOLGA4 and TGN38. Fluorescence images for fixed cells were taken with confocal fluorescence microscope (SpinSR10; Olympus) and (LSM800; ZEISS).
3
Immunofluorescence Analysis of Epithelial-Mesenchymal Transition Markers
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The indicated cells were seeded on the confocal dishes overnight, washed with 1 × PBS three times, fixed in 4% paraformaldehyde for 30 min, blocked with 1% bovine serum albumin for 1 h and incubated with antibodies against E-Cadherin (1:150, Proteintech, 20874-1-AP), N-Cadherin (1:150, Proteintech, 22018-1-AP), and Vimentin (1:150, Proteintech, 10366-1-AP) antibodies for 1 h at room temperature (RT). Then, the cells were incubated with Alexa Fluor 594-conjugated secondary antibodies (Molecular Probes, USA) for 1 h at RT and subsequently mounted with the DAPI-containing mounting medium (Beyotime, Shanghai, China). The images were taken with a laser scanning confocal microscope (Leica Microsystems AG).
4
Immunofluorescence Localization of Claudin-1
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Localization of Claudin‐1, one representative of tight junction proteins in intestine in each group, was determined by immunofluorescence microscopy. First, cryosection of intestine tissues was washed with PBS. For Claudin‐1 staining, cryosection of intestine tissues was fixed with 2% (v/v) formaldehyde in PBS for 30 min and incubated with 1% (v/v) Triton X‐100 in PBS 3 times for 5 min each to permeabilize cells and then washed and blocked for 30 min with 1% (w/v) bovine serum albumin (BSA) in PBS. Cryosection was then incubated with primary antibody against rabbit Claudin‐1 (1:100) overnight at 4°C. Cryosection was then washed again with PBS and incubated with the secondary antibody, Alexa Fluor 488 goat anti‐rabbit (1:100) (Proteintech). Cryosection was then washed again with PBS, and the permeable support membrane was then mounted side up between a slide and coverslip with a DAPI‐containing mounting medium (Beyotime Biotechnology). Microscopic images of the mounted membranes were taken on a Zeiss LSM 510 Meta Confocal Laser Scanning Microscope (Leica, DFC450C). Scan areas were chosen based on intact tissue structures, avoiding the edges of the square cutouts.
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