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Storm 860 system

Manufactured by GE Healthcare
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The STORM 860 system is a high-performance biomolecular imager designed for life science research. It utilizes a unique combination of laser illumination and advanced image capture technology to enable researchers to visualize and analyze a wide range of biomolecular samples, such as fluorescently labeled proteins, nucleic acids, and other biological molecules.

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Lab products found in correlation

C16 pi, by Echelon Biosciences (2 mentions) Pi3k p110γ, by Merck Group (2 mentions) Kinase buffer, by Cell Signaling Technology (1 mentions)

Close spelling variants of this product

Storm 860 PhosphorImager system Storm 860 Gel and Blot Imaging System Storm 860 Imaging System Storm 860

5 protocols using storm 860 system

1

Quantifying PI4KIIIα Enzymatic Activity

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The relative activity of the different PI4KIIIα preparations was assayed as follows. Liposomes containing C16-PI (Echelon Biosciences) were generated by sonication of a 1 mg/mL solution in kinase buffer (100 mM Tris-HCl pH 7.5, 50 mM EGTA, 100 mM MgCl2)) and incubated (at 80 μM) in a 50 μL reaction with 400 ng of PI4KIIIα or PI4KIIIα complex (equal amounts of PI4KIIIα in each sample was ensured by the use of both the BCA assay and SDS-PAGE/Coomassie staining), γ32P-labeled ATP (10 μCi), and cold ATP (50 μM) in kinase buffer for 5 min at 37 °C. The reaction was quenched by the addition of 700 μL of 2:1 chloroform:methanol containing 10 μg/mL Folch fraction (brain phosphoinositides) and 400 μL of 0.1 M HCl. The organic extracts were dried, resuspended in a small amount of 1:1 chloroform:methanol, and equal amounts were analyzed by thin-layer chromatography (mobile phase is 14:32:24:30:64 water:acetic acid:methanol:acetone:chloroform). PI4P was identified by comparison with PI3P generated in a parallel reaction using PI3K p110γ (Sigma) and quantified by autoradiography using a STORM 860 system (Molecular Dynamics).
Baskin J.M., Wu X., Christiano R., Oh M.S., Schauder C.M., Gazzerro E., Messa M., Baldassari S., Assereto S., Biancheri R., Zara F., Minetti C., Raimondi A., Simons M., Walther T.C., Reinisch K.M, & De Camilli P. (2015). The leukodystrophy protein FAM126A/Hyccin regulates PI4P synthesis at the plasma membrane. Nature cell biology, 18(1), 132-138.
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2

Steroid Metabolism in LAPC-4 Cells

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LAPC-4 cells were plated in 24-well culture at 20,000 cells per well. After incubation for 2 days, 50 nM of [14C]-4-androstene-3,17-dione (Perkin Elmer Life Sciences Inc., Boston, MA, USA) and 10 μL of RM-532-105 dissolved in DMSO and culture medium were added. The final DMSO concentration in each was adjusted to 0.05%. After 3 and 5 days, the culture medium was removed from wells and steroids were extracted with diethyl ether. After evaporating the organic phase to dryness with a nitrogen stream, residue was dissolved in dichloromethane, dropped on silica gel 60 F254 thin layer chromatography (TLC) plates (EMD Chemicals Inc., Gibbstown, NJ, USA) and eluted with a mixture of toluene/acetone (4:1). [14C]-labeled steroids (4-dione, A-dione, DHT/ADT, epi-ADT and T) were identified by comparison with reference steroids and quantified using the Storm 860 System (Molecular Dynamics, Sunnyvale, CA, USA). The relative abundance of each steroid was calculated and expressed in percentage.
Kenmogne L.C., Roy J., Maltais R., Rouleau M., Neveu B., Pouliot F, & Poirier D. (2017). Investigation of the In Vitro and In Vivo efficiency of RM-532-105, a 17β-hydroxysteroid dehydrogenase type 3 inhibitor, in LAPC-4 prostate cancer cell and tumor models. PLoS ONE, 12(2), e0171871.
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3

Quantifying PI4KIIIα Enzymatic Activity

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The relative activity of the different PI4KIIIα preparations was assayed as follows. Liposomes containing C16-PI (Echelon Biosciences) were generated by sonication of a 1 mg/mL solution in kinase buffer (100 mM Tris-HCl pH 7.5, 50 mM EGTA, 100 mM MgCl2)) and incubated (at 80 μM) in a 50 μL reaction with 400 ng of PI4KIIIα or PI4KIIIα complex (equal amounts of PI4KIIIα in each sample was ensured by the use of both the BCA assay and SDS-PAGE/Coomassie staining), γ32P-labeled ATP (10 μCi), and cold ATP (50 μM) in kinase buffer for 5 min at 37 °C. The reaction was quenched by the addition of 700 μL of 2:1 chloroform:methanol containing 10 μg/mL Folch fraction (brain phosphoinositides) and 400 μL of 0.1 M HCl. The organic extracts were dried, resuspended in a small amount of 1:1 chloroform:methanol, and equal amounts were analyzed by thin-layer chromatography (mobile phase is 14:32:24:30:64 water:acetic acid:methanol:acetone:chloroform). PI4P was identified by comparison with PI3P generated in a parallel reaction using PI3K p110γ (Sigma) and quantified by autoradiography using a STORM 860 system (Molecular Dynamics).
Baskin J.M., Wu X., Christiano R., Oh M.S., Schauder C.M., Gazzerro E., Messa M., Baldassari S., Assereto S., Biancheri R., Zara F., Minetti C., Raimondi A., Simons M., Walther T.C., Reinisch K.M, & De Camilli P. (2015). The leukodystrophy protein FAM126A/Hyccin regulates PI4P synthesis at the plasma membrane. Nature cell biology, 18(1), 132-138.
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4

Kinase Assay for PI4KA Activity

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The kinase assay was performed as previously described (Baskin, et al., 2016 (link)) with modifications. Briefly, HA-tagged PI4KA protein was immunoprecipitated from HEK293T cells expressing PI4KA-HA and reconstituted in the kinase buffer (9802, Cell signaling). Liposomes containing 2μg C16-PtdIns (Echelon Biosciences) generated by sonication in the kinase buffer, γ33P-labelled ATP (15 μCi), and cold ATP (50 μM) were mixed with PI4KA-HA protein and incubated with or without recombinant SRGAP2 protein for 30 min at 37°C. The reaction was quenched by the addition of 700 μl of 2:1 chloroform/methanol containing 10 μg/ml Folch fraction (brain phosphoinositides) and 400 μl of 0.1M HCl. The organic extracts were dried, resuspended in a small amount of 1:1 chloroform/methanol, and analyzed by thin-layer chromatography (mobile phase is 14:32:24:30:64 water/acetic acid/methanol/acetone/chloroform). PtdIns(4)P was quantified by autoradiography using a STORM 860 system (Molecular Dynamics).
Ren C., Yuan Q., Braun M., Zhang X., Petri B., Zhang J., Kim D., Guez-Haddad J., Xue W., Pan W., Fan R., Kubes P., Sun Z., Opatowsky Y., Polleux F., Karatekin E., Tang W, & Wu D. (2019). Leukocyte cytoskeleton polarization is initiated by plasma membrane curvature from cell attachment. Developmental cell, 49(2), 206-219.e7.
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5

DHEA Metabolism in Breast Cancer Cells

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T-47D cells (3000, 6000, 12,000 or 24,000) were plated in 24-well culture. After incubation for 1 day, 10 μL of a 50 μM [14C]-DHEA solution (Perkin Elmer, Boston, MA, USA) in culture medium (990 μL) were added. After 1, 2, 3, 6 or 8 days, the culture medium was recovered from the wells and steroids were extracted with diethyl ether. After evaporating the organic phase to dryness with a nitrogen stream, the residue was dissolved in dichloromethane, dropped on silica gel 60F254 thin-layer chromatography (TLC) plates (Milipore-Sigma, Oakville, ON, Canada) and eluted with a mixture of toluene:acetone (4:1), [14C]-labeled steroids (DHEA, 5-diol, 4-dione, T, DHT, E1, E2, ADT and 3β-diol) were identified by comparison with reference steroids and quantified using a Storm 860 System (Molecular Dynamics, Sunnyvale, CA, USA). The relative abundance of each steroid was calculated and expressed in percentage. In one experiment, steroids (DHEA, 5-diol, 4-dione, DHT and E2) were quantified by gas chromatography–mass spectrometry (GC-MS/MS) analysis according to an established procedure developed at the CHU de Québec (CHUL)-Research Center and previously published [65 (link)].
Poirier D., Roy J, & Maltais R. (2021). A Targeted-Covalent Inhibitor of 17β-HSD1 Blocks Two Estrogen-Biosynthesis Pathways: In Vitro (Metabolism) and In Vivo (Xenograft) Studies in T-47D Breast Cancer Models. Cancers, 13(8), 1841.
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