BeWo cell syncytialisation was induced by forskolin as previously described [9 (link)]. Briefly, BeWo cells were treated with 60 μM forskolin (Cell Signaling Technology, #3828s) for 48 hours to induce cell fusion and the culture medium with 60 μM forskolin was replaced every 24 h. After treatments, BeWo cells were incubated with Hoechst (Invitrogen™, #H3570, 1:1500) and various fluorescent membrane dyes: 2 μM Di-8-ANEPPS (Invitrogen™, # D3167), 5.0 μg/mL Wheat Germ Agglutinin-Alexa Fluor™−488 Conjugate (Invitrogen™, # W11261), 1:200 CellBrite™-Orange Cytoplasmic Membrane Dye (Biotium, #30022), and 50 μg/mL Concanavalin A-Alexa Fluor™−647 Conjugate (Invitrogen, #C21421) in 5% CO2 at 37 °C for 15–20 minutes. Di-8-ANEPPS stained cells can be directly imaged without extra rinsing steps. The other membrane markers need to be washed twice with the dye-free medium at room temperature.
Cellbrite orange cytoplasmic membrane dye
Manufactured by Biotium
CellBrite™-Orange Cytoplasmic Membrane Dye is a fluorescent dye that selectively labels the cytoplasmic membranes of cells. It can be used to visualize cell morphology and distribution in various cell-based applications.
Automatically generated - may contain errors
Lab products found in correlation
Forskolin, by Thermo Fisher Scientific (1 mentions)
Hoechst, by Thermo Fisher Scientific (1 mentions)
Di 8 anepps, by Thermo Fisher Scientific (1 mentions)
Forskolin, by Cell Signaling Technology (1 mentions)
Concanavalin a alexa fluor 647 conjugate, by Thermo Fisher Scientific (1 mentions)
Matrigel matrix growth factor reduced, by BD (1 mentions)
2 protocols using cellbrite orange cytoplasmic membrane dye
1
Forskolin-Induced BeWo Cell Syncytialisation
2
In vivo Skin Regeneration Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
hDP cells (passage 2 or 3; average 2.6 × 105), iMCs or iDPSCs (average 3.6 × 105) were stained with CellBrite Orange Cytoplasmic Membrane Dye (Biotium), mixed with Matrigel Matrix Growth Factor Reduced (BD Biosciences) and placed onto thin silicone sheets. Subsequently, cultured human adult KCs (average 1.5 × 105) in Matrigel were placed on top. Then, composites were fully covered with cultured human fibroblasts (1.2 × 104/μl) in Matrigel. Final composites were transplanted into the dorsal region of anesthetised 8-week-old female C.B-17/IcrHsd-Prkdcscid mice (Japan SLC). After 5–6 weeks, grafts were harvested and microdissected using watchmaker’s forceps and fine needles under light microscopy. All animal procedures were performed in accordance with the guidelines of the Science Council of Japan and approved by the Keio University Institutional Animal Care and Use Committee.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!