Log In Sign up
The largest database of trusted experimental protocols

Cyclopamine

Manufactured by LGC
Visit Supplier
Sourced in United States

Cyclopamine is a naturally occurring steroidal alkaloid compound. It functions as a potent inhibitor of the Hedgehog signaling pathway, a key regulator of embryonic development and tissue homeostasis.

Automatically generated - may contain errors

Lab products found in correlation

20 s ohc, by Steraloids (2 mentions) Sant 1, by Merck Group (2 mentions) Recombinant shh n protein, by R&D Systems (1 mentions) Cell count reagent sf, by Nacalai Tesque (1 mentions) Rhshh, by R&D Systems (1 mentions) Purmorphamine, by Merck Group (1 mentions) Tamoxifen, by R&D Systems (1 mentions) Fibroblast medium, by ScienCell (1 mentions) Fk506, by Cayman Chemical (1 mentions) Tomatidine, by Merck Group (1 mentions) Horseradish peroxidase conjugated secondary antibody, by Cell Signaling Technology (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Propidium iodide, by Merck Group (1 mentions) Fetal bovine serum (fbs), by Corning (1 mentions) Heparin acrylic, by Merck Group (1 mentions) Heparin, by LGC (1 mentions) Affi gel blue, by Bio-Rad (1 mentions)

15 protocols using cyclopamine

1

Cyclopamine Treatment of Sea Urchin Embryos

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cyclopamine treatments were performed as described previously (Tarazona et al., 2016 (link)) with the following modifications; stage 16 embryos were treated with 10 μM Cyclopamine (C988400, Toronto Research Chemicals) for 2 days, then washed thoroughly ten times with FASW. Embryos were then washed five more times every hour and one time every day before collecting the embryos for SEM. Control embryos were treated with 0.1% DMSO and then washed as described above.
Tarazona O.A., Lopez D.H., Slota L.A, & Cohn M.J. (2019). Evolution of limb development in cephalopod mollusks. eLife, 8, e43828.
+ Open protocol
+ Expand
2

Molecular Components of the Hedgehog Signaling Pathway

Cited 2 times
Check if the same lab product or an alternative is used in the 5 most similar protocols
NIH 3T3 and 293T cells were obtained from ATCC, Smo−/− fibroblasts have been described previously33 (link) and were originally obtained from Drs. James Chen and Philip Beachy, SAG was from Enzo Life Sciences, Cyclopamine from Toronto Research Chemicals, SANT-1 from EMD Millipore, 20(S)-OHC from Steraloids. The synthesis of 20(R)-OHC7 (link) and 20(S)-amine coupled beads for SMO binding assays has been described in detail previously10 (link). Rabbit polyclonal antibodies against SMO, PTCH1 and SUFU have been described previously34 (link),35 (link), and the mouse monoclonal antibody against GLI1 was obtained from Cell Signaling Technologies (Cat#L42B10). SHH-containing conditioned media was made from 293T cells transfected with the N-terminal signalling domain of SHH and used at saturating concentrations (dilution of 1:4)2 (link).
Byrne E.F., Sircar R., Miller P.S., Hedger G., Luchetti G., Nachtergaele S., Tully M.D., Mydock-McGrane L., Covey D.F., Rambo R.P., Sansom M.S., Newstead S., Rohatgi R, & Siebold C. (2016). Structural basis for Smoothened regulation by its extracellular domains. Nature, 535(7613), 517-522.
+ Open protocol
+ Expand
3

Intravitreal Injections for Retinal Regeneration

Cited 2 times
Check if the same lab product or an alternative is used in the 5 most similar protocols
Intravitreal injections were performed as previously described (Qin et al., 2011 (link); Thomas et al., 2016 (link)). Fish were anesthetized and a small incision was made in the cornea using a Safety Sideport Straight Knife (15°; Beaver-Vistec International). A 33-gauge blunt-end Hamilton Syringe was used to inject 0.5–0.75 microliters of solution. Ouabain injections (10 µM) were performed to damage all retinal neurons as previously described (Fimbel et al., 2007 (link); Sherpa et al., 2014 (link)). Gain- and loss-of-function studies utilized 1X PBS or 1% EtOH for control solutions, and recombinant SHH-N protein (100 µg/mL in 1X PBS; R&D Systems) or cyclopamine (100 µM in 1% EtOH; Toronto Research Chemicals). Light damaged zebrafish were injected beginning at 2 days prior to light onset (− 2dpL) and continued daily through 2 dpL, with amaximum of 5 total injections (Suppl. Fig. 1A). Ouabain damaged retinas were injected beginning at 3 dpi and continued through 10 dpi, with a maximum of 8 total injections (Suppl Fig 1B).
Thomas J.L., Morgan G.W., Dolinski K.M, & Thummel R. (2017). Characterization of the Pleiotropic Roles of Sonic Hedgehog during Retinal Regeneration in Adult Zebrafish. Experimental eye research, 166, 106-115.
+ Open protocol
+ Expand
4

Evaluating Hedgehog Pathway Regulation

Check if the same lab product or an alternative is used in the 5 most similar protocols
GBd15 and TGBC2TKB cells were seeded onto 96-well plates at a density of 5000 cells/well and incubated with recombinant human Shh (rhShh; R&D Systems, Minneapolis, MN, USA) or the Smo inhibitor cyclopamine (Toronto Research Chemicals, North York, Canada) for 24, 48, or 72 h. Cell proliferation was determined by the absorbance at 492 nm (ref. 620 nm) using Cell Count Reagent SF (Nacalai Tesque, Kyoto, Japan). Forty-eight hours after Smo, MMP-2, and MMP-9 siRNA transfection, the cells were reseeded onto 96-well plates and the proliferation rates were measured.
Matsushita S., Onishi H., Nakano K., Nagamatsu I., Imaizumi A., Hattori M., Oda Y., Tanaka M, & Katano M. (2014). Hedgehog signaling pathway is a potential therapeutic target for gallbladder cancer. Cancer Science, 105(3), 272-280.
+ Open protocol
+ Expand
5

Molecular Components of the Hedgehog Signaling Pathway

Cited 2 times
Check if the same lab product or an alternative is used in the 5 most similar protocols
NIH 3T3 and 293T cells were obtained from ATCC, Smo−/− fibroblasts have been described previously33 (link) and were originally obtained from Drs. James Chen and Philip Beachy, SAG was from Enzo Life Sciences, Cyclopamine from Toronto Research Chemicals, SANT-1 from EMD Millipore, 20(S)-OHC from Steraloids. The synthesis of 20(R)-OHC7 (link) and 20(S)-amine coupled beads for SMO binding assays has been described in detail previously10 (link). Rabbit polyclonal antibodies against SMO, PTCH1 and SUFU have been described previously34 (link),35 (link), and the mouse monoclonal antibody against GLI1 was obtained from Cell Signaling Technologies (Cat#L42B10). SHH-containing conditioned media was made from 293T cells transfected with the N-terminal signalling domain of SHH and used at saturating concentrations (dilution of 1:4)2 (link).
Byrne E.F., Sircar R., Miller P.S., Hedger G., Luchetti G., Nachtergaele S., Tully M.D., Mydock-McGrane L., Covey D.F., Rambo R.P., Sansom M.S., Newstead S., Rohatgi R, & Siebold C. (2016). Structural basis for Smoothened regulation by its extracellular domains. Nature, 535(7613), 517-522.
+ Open protocol
+ Expand
6

Quantifying Smo Localization in MEFs

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
Mouse embryonic fibroblasts (MEFs) were isolated and immortalized as previously described [53 ]. Control and Smocbb MEFs were grown on coverslips at a density of 0.5 × 106 cells/mL and treated for 24 h with 0.5% fetal bovine serum (FBS) Shh-conditioned medium [54 (link)], 0.5% FBS medium containing 100 or 400 nM SAG (Millipore), 0.5% FBS medium containing 5 uM cyclopamine (Toronto Research Chemicals), or 0.5% FBS DMEM. Ten images were taken of each coverslip and scored by two independent reviewers blinded to genotype. Smo localization in cilia was categorized as full, partial, or negative (Figure 6A).
Gigante E.D., Long A.B., Ben-Ami J, & Caspary T. (2018). Hypomorphic Smo mutant with inefficient ciliary enrichment disrupts the highest level of vertebrate Hedgehog response. Developmental biology, 437(2), 152-162.
+ Open protocol
+ Expand
7

Modulating Sonic Hedgehog Signaling in Zebrafish

Check if the same lab product or an alternative is used in the 5 most similar protocols
Sonic hedgehog signaling was modulated with either an agonist (purmorphamine, Sigma-Aldrich, St. Louis, MO, USA) or an antagonist (cyclopamine, Toronto Research Chemicals, Toronto, ON, Canada). purmorphamine: Undamaged fish were IP-injected with 40 µL of 10 µM purmorphamine using a 30-gauge injection needle attached to a 1 mL syringe every 12 h for 48 h (0, 12, 24, 36, and 48 h). To assess cell proliferation in undamaged control and purmorphamine treated brains, EdU was coinjected (as described above) at the 48 h timepoint. Brains were collected 12 h later (60 h after the first purmorphamine injection) and prepared for EdU detection and immunohistochemistry as described above. cyclopamine: Fish exposed to sTBI were IP-injected with 40 µL of 2 mM cyclopamine at 4, 12, 24, 36 hpi using a 30-gauge injection needle attached to a 1 mL syringe and coinjected with EdU at 48 hpi. Brains were collected at 60 hpi and prepared for EdU detection and immunohistochemistry as described above.
Hentig J., Cloghessy K., Lahne M., Jung Y.J., Petersen R.A., Morris A.C, & Hyde D.R. (2021). Zebrafish Blunt-Force TBI Induces Heterogenous Injury Pathologies That Mimic Human TBI and Responds with Sonic Hedgehog-Dependent Cell Proliferation across the Neuroaxis. Biomedicines, 9(8), 861.
+ Open protocol
+ Expand
8

Compound Preparation for Cell Studies

Check if the same lab product or an alternative is used in the 5 most similar protocols
If not otherwise stated all compounds were obtained from Sigma-Aldrich. For in vitro studies 5 μM cyclopamine (Toronto Research Chemicals Inc.) dissolved in ethanol (EtOH), 10 μM tamoxifen in DMSO and 1 μg/ml recombinant murine Shh-N (rShh-N; R&D systems) in HD-buffer were used. tamoxifen solution for in vivo use was prepared as described (see below)23 (link).
Pyczek J., Buslei R., Schult D., Hölsken A., Buchfelder M., Heß I., Hahn H, & Uhmann A. (2016). Hedgehog signaling activation induces stem cell proliferation and hormone release in the adult pituitary gland. Scientific Reports, 6, 24928.
+ Open protocol
+ Expand
9

Modulating Bladder Cell Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols
Primary human bladder urothelial cells (ScienCell 4320) and bladder stromal fibroblasts (ScienCell 4330) were cultured in Urothelial Cell Medium (ScienCell 4321) and Fibroblast Medium (ScienCell 2301) respectively to sub-confluency. Cells were then starved for 12 hours, followed by addition of Shh conditioned medium in the absence or in the presence of cyclopamine (3uM, Toronto Research Chemicals), or treated with the Shh pathway agonist SAG (200 nM, Millipore) for 24 hours. In separate experiments, primary human bladder urothelial cells (ScienCell 4320) or J82 human urothelial carcinoma cells (ATCC HTB1) were pre-treated with 120 nM LDN-193189 or DMSO control (Selleckchem S2618) for 30 min prior to treatment with 20ng/ml FK506 (Cayman Chemical) or treated with a DMSO control for 12 hours. Following treatment, RNA was extracted using the Trizol reagent.
Shin K., Lim A., Zhao C., Sahoo D., Pan Y., Spiekerkoetter E., Liao J.C, & Beachy P.A. (2014). Hedgehog signaling restrains bladder cancer progression by eliciting stromal production of urothelial differentiation factors. Cancer cell, 26(4), 521-533.
+ Open protocol
+ Expand
10

Cyclopamine Embryo Developmental Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols
Cyclopamine was obtained from Toronto Research Chemicals and added to embryos from a 10 mM stock prepared in ethanol. Dechorionated embryos (20 animals per group) were treated in E3 medium containing 20–60 µM Cyclopamine from 32–34 hpf and kept in Cyclopamine solution until the desired time point. Embryos were then fixed in 4% paraformaldehyde for staining.
Boyd P.J., Cunliffe V.T., Roy S, & Wood J.D. (2015). Sonic hedgehog functions upstream of disrupted-in-schizophrenia 1 (disc1): implications for mental illness. Biology Open, 4(10), 1336-1343.
+ Open protocol
+ Expand

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

Sign up now

Revolutionizing how scientists
search and build protocols!