Verapamil
Manufactured by Spirochrome
Sourced in Switzerland
Verapamil is a calcium channel blocker used in the laboratory setting. It functions by inhibiting the movement of calcium ions across cell membranes, which can affect various cellular processes.
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Lab products found in correlation
Sir tubulin, by Spirochrome (3 mentions)
Lsm 880, by Zeiss (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Sir dna, by Spirochrome (1 mentions)
Cage incubator, by Okolab (1 mentions)
Sodium arsenite, by Merck Group (1 mentions)
Oxyrase, by Merck Group (1 mentions)
Leibovitz s l 15 medium, by Thermo Fisher Scientific (1 mentions)
Ds qi2 camera, by Nikon (1 mentions)
Collagen 1, by BD (1 mentions)
Sir actin, by Spirochrome (1 mentions)
Endothelial cell growth supplement (ecgs), by Merck Group (1 mentions)
M199 medium, by Thermo Fisher Scientific (1 mentions)
Ti wide field microscope, by Nikon (1 mentions)
Coolsnap hq camera, by Teledyne (1 mentions)
L ascorbic acid, by Merck Group (1 mentions)
Hydrocortisone, by Merck Group (1 mentions)
Heparin sodium salt, by Merck Group (1 mentions)
Lysotracker red dnd 99, by Thermo Fisher Scientific (1 mentions)
Non essential amino acids (neaa), by Thermo Fisher Scientific (1 mentions)
7 protocols using verapamil
1
Live Cell DNA Staining and Imaging
2
Visualizing Cytoskeleton Dynamics in U2OS Cells
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Single U2OS cells stably expressing GFP-tagged G3BP1 (Pelkmans Lab64 (link)) were plated on patterned coverslips following the protocol mentioned above. After spreading for 3–4 h, the cells were switched into media containing 100 nM SiR-tubulin (Spirochrome) and 10 μM verapamil (Spirochrome) and incubated in the dye for at least 4 h. The cells were imaged at 37 °C in imaging media containing Leibovitz’s L-15 medium (ThermoFisher Scientific) with 10% fetal bovine serum, 30 μl ml–1 Oxyrase (Sigma-Aldrich) and 10 mM lactic acid, with 0.5 mM sodium arsenite (Sigma-Aldrich). Imaging was done on a Nikon Ti2 Eclipse epifluorescent microscope with a cage incubator (Okolab), using ×100 oil objective with numerical aperture (NA) of 1.49 and a Nikon DS-Qi2 camera.
3
Actin and Nuclear Cytometry in Cells
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Triangle patterned coverslips (4Dcell) were coated with 1.5 µg/cm² collagen I (BD-Biosciences) for 1 h and blocked with 1% BSA in PBS for 1 h at room temperature. 1x10 4 cells were seeded, and after 4 h, cells were incubated overnight with 0.5 µM SiR-actin and 10 µM Verapamil (Spirochrome). Cells were fixed with 4% paraformaldehyde (Merck) for 10 min at room temperature. Nuclei were stained with Hoechst 33258 and coverslips were mounted in Fluoromount as described above.
4
Actin Dynamics Imaging in Tissue Culture
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Standard α-MEM was exchanged with α-MEM supplemented with 100 nM SiR-actin (633 nm, Spirochrome, Switzerland) and 10 μM verapamil (Spirochrome, Switzerland) 24 hours before imaging and then incubated at 37°C in a humidified atmosphere with 5% CO2. Samples were imaged after 5, 10, 15, and 32 days of tissue culture with a Leica SP8 confocal microscope using a 25× 0.95-NA water immersion objective lens (Leica Fluotar VISIR) and a helium-neon laser at 633 nm. After each imaging cycle, medium containing SiR-actin was exchanged with standard α-MEM to reduce prolonged SiR-actin exposure.
5
Visualizing Crenolanib's Effects on Mitosis
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To visualise the effect of crenolanib on mitosis, live-cell imaging was performed for 24 h at 37 °C on a Ti widefield microscope (Nikon) equipped with an environmental chamber (5% CO2) using a 60 × 1.3 NA oil objective, a Cy5 filter, a CoolSNAP HQ camera (Roper Scientific, Vianen, Netherlands) at a sampling rate of 3 min, recording at each time point 9 z-stacks separated by 2 µm. ECRF24 were maintained as described above. HUVECs were maintained in M199 medium (Thermo Fisher Scientific) supplemented with 10% fetal calf serum (FCS), 1% penicillin/streptomycin (Thermo Fisher Scientific), 1% endothelial cell growth supplement (Millipore), 0.1 mg/mL heparin sodium salt (Sigma), 0.1 µM hydrocortisone (Sigma), and 10 µg/mL L-ascorbic acid (Sigma). The day before, the cells were seeded in an Ibidi µ-Slide 8 Well culture plate (Vitaris, Baar, Switzerland). 4 h prior to the start of imaging, 50 nM SiR-Tubulin (SpiroChrome AG, Switzerland) was added on the cells to stain microtubules in addition with 10 µM Verapamil (SpiroChrome AG, Stein am Rein, Switzerland) to keep the dye inside the cells. None of these compounds affected mitosis, nor the efficacy of crenolanib. Crenolanib was added just before the start of image acquisition. Time-lapse movies were analysed using NIS Elements AR Software.
6
NK Cell Cytotoxicity Visualization Protocol
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5×104 SKBR3 cells were seeded in a µ-Slide 8 Well Glass Bottom overnight (Ibidi GmbH, Martinsried, Germany). 105 NK cells were washed once with phenol red-free RPMI-1640 medium, supplemented 20 mM HEPES (Sigma Aldrich), 2 mM L-glutamine, and non-essential amino acids (Life Technologies) and stained with SiR-tubulin and verapamil (Spirochrome) (500 nM and 10 µM, respectively) for 1 hour at 37°C. LysoTracker Red DND-99 (Life technologies) was then added to NK cells (10 µM) for another 30 min at 37°C. Following incubation, NK cells were washed three times and treated or not with 100 ng/mL CHI3L1 for 1 hour. In parallel, SKBR3 targets were treated with 10 µg/mL Trastuzumab for 20 min. After mounting the slide with the SKBR3 target cells, the medium containing Trastuzumab was removed before the addition of NK cells. Eight fields per condition were recorded for 2 hours with 180 s intervals. Live imaging was performed at 37°C using environmental chambers on a Leica SD AF Spinning Disc Confocal (Leica-microsystems).
7
Cell Staining with SiR-DNA and Verapamil
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One hour prior to the experiment, the media on cells in the magnetic microarray was replaced with culture media containing 1µM SiR-DNA and 10µM verapamil (Spirochrome). Subsequently, media was replaced with normal culture media.
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