Enlightning

Manufactured by PerkinElmer

Sourced in United States

Enlightning is a compact and versatile spectrophotometer designed for accurate and reliable analysis of various samples. It features a high-resolution display, intuitive software, and advanced optics to deliver precise measurements across a wide range of applications.

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Lab products found in correlation

T7 scribe standard rna ivt kit, by CellScript (1 mentions) Anti rabbit igg fitc antibody, by Merck Group (1 mentions) Lipofectamine ltx and plus reagent, by Thermo Fisher Scientific (1 mentions) Transdirect insect cell, by Shimadzu (1 mentions) 3h leucine, by GE Healthcare (1 mentions) Ecl prime western blotting detection reagent, by GE Healthcare (1 mentions) Mitotracker red cmxros, by Thermo Fisher Scientific (1 mentions) Anti flag monoclonal antibody, by Merck Group (1 mentions) Protein g hrp conjugate, by Bio-Rad (1 mentions) Hoechst 33342, by Thermo Fisher Scientific (1 mentions) X ray film, by Kodak (1 mentions) 3h adomet, by PerkinElmer (1 mentions) Bis tris gel, by Thermo Fisher Scientific (1 mentions) Biomax ms film, by Kodak (1 mentions) Gel dryer 583, by Bio-Rad (1 mentions) Mini protean tgx stain free gel, by Bio-Rad (1 mentions)

5 protocols using enlightning

Kinetic Analysis of MLL1 Automethylation

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Apparent K_m values for AdoMet in MLL1 automethylation reactions were determined by incubating wild type or variant MLL1 at a concentration of 14 µM with various concentrations of [³H]-AdoMet (0–50 µM) at 15°C. The samples were quenched with SDS loading buffer at one and three-hour time points and separated by 4–12% Bis-Tris SDS-PAGE and stained with Coomassie brilliant blue. Gels were soaked in an enhancer solution (Enlightning, Perkin Elmer, Inc.) for 30 minutes then dried and exposed to film for 7 days at −80°C. Bands were quantified by densitometry using Image J^{66 (link)}, and relative intensity values were plotted as a function of time and fitted by linear regression to determine methylation rates. Methylation rates were then plotted as a function of AdoMet concentration and fitted by nonlinear least-squares regression to the Michaelis-Menton Equation 1. We note that that all variants tested displayed a pattern of substrate inhibition above 20 µM AdoMet, therefore apparent K_m values are reported from the data fitted over the concentration range from 0–20 µM AdoMet.

v = v_{max} \times \frac{[AdoMet]}{K_{m} + [AdoMet]}

Shinsky S.A., Hu M., Vought V.E., Ng S.B., Bamshad M.J., Shendure J, & Cosgrove M.S. (2014). A non-active site SET domain surface crucial for the interaction of MLL1 and the RbBP5-ASH2L heterodimer within MLL family core complexes. Journal of molecular biology, 426(12), 2283-2299.

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SDS-PAGE Protein Visualization

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The radiolabeled proteins were resolved by 12.5% SDS-PAGE, then the gel was fixed and soaked in ENLIGHTNING (PerkinElmer) for 20 min. Thereafter, the gel was dried under vacuum and exposed to X-ray film for an appropriate period.

Takamitsu E., Otsuka M., Haebara T., Yano M., Matsuzaki K., Kobuchi H., Moriya K, & Utsumi T. (2015). Identification of Human N-Myristoylated Proteins from Human Complementary DNA Resources by Cell-Free and Cellular Metabolic Labeling Analyses. PLoS ONE, 10(8), e0136360.

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Insect Cell-Free Protein Synthesis Protocol

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Transdirect insect cell, an insect cell-free protein synthesis system, was obtained from Shimadzu (Kyoto, Japan). Human cDNAs (Flexi ORF clones) were purchased from Promega (Madison, WI, USA). [³H]leucine, [³H]myristic acid, and ECL prime western blotting detection reagent were from GE Healthcare (Buckinghamshire, UK). ENLIGHTNING was from PerkinElmer (Waltham, MA, USA). T7-Scribe standard RNA IVT kit was from CELLSCRIPT (Madison, WI, USA). The dye terminator cycle sequencing kit, Lipofectamine LTX and Plus reagent, MitoTracker Red CMXRos and Hoechst 33342 were from Life Technologies Corporation (Carlsbad, CA, USA). Anti-FLAG monoclonal antibody, anti-SAMM50 monoclonal antibody (WH0025813), anti-SAMM50 polyclonal antibody (HPA034537) and anti-Rabbit IgG-FITC antibody were from Sigma (St. Louis, MO, USA). Protein G-HRP conjugate was from Bio-Rad (Hercules, CA, USA). ImmunoStar LD was from Wako Pure Chemical (Osaka, Japan). X-ray film was from Eastman Kodak (Rochester, NY, USA). The other reagents used were from Wako Pure Chemical (Osaka, Japan), Daiichi Pure Chemicals (Tokyo, Japan) or Seikagaku Kogyo (Tokyo, Japan) and were of analytical or DNA grade.

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Enzymatic Assay of MLL3 SET Domain

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Purified MLL3 SET domains (wild type and mutant) alone (5 μm) or assembled with a stoichiometric amount of WRAD/RAD (5 μm) were incubated with 1.5 μCi of [³H]AdoMet (PerkinElmer Life Sciences, specific activity 78 Ci/mmol) at 15 °C for 3 h and then incubated on ice for 1 h. Half the volume of each sample was quenched with SDS loading buffer, and the other half was exposed to UV light (254 nm) in a Stratalinker oven at a distance of ∼15 cm for 30 min on ice. The UV-exposed samples were quenched with SDS loading buffer, and all samples were separated by SDS-PAGE using a 4–12% BisTris gel (Invitrogen) run at 200 V for 30 min. The gels were soaked in enhancer solution for 30 min (Enlightning, PerkinElmer Life Sciences) and then dried for 2.5 h at 72 °C under constant vacuum. The dried gels were exposed to film (Kodak Biomax MS Film) for 72 h at −80 °C.

Shinsky S.A, & Cosgrove M.S. (2015). Unique Role of the WD-40 Repeat Protein 5 (WDR5) Subunit within the Mixed Lineage Leukemia 3 (MLL3) Histone Methyltransferase Complex. The Journal of Biological Chemistry, 290(43), 25819-25833.

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Quantifying Viral Protein Synthesis in A549 Cells

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A549 cells were infected with 4.43 × 10⁸ PFU/ml of CVB3. The virus was bound on ice for 1 h, after which the excess virus was washed with PBS. IDPN (1.5%) in DMEM supplemented with 1% FBS was added after ice binding. After IDPN was added, it was present at all steps until the end of the experiment. The infection was allowed to proceed at 37°C for 4 h, after which the low methionine/cysteine medium supplemented with dialyzed 1% FBS was added to cells. After 30 min, 500 μCi/ml of [³⁵S]methionine-cysteine was added before a 1-h pulse. Samples were run at 4 to 20% Mini-Protean TGX Stain-Free gel (Bio-Rad), after which the gel was fixed with 30% methanol, 10% acetic acid for 30 min. The gel then was treated with an autoradiography enhancer (Enlightning; PerkinElmer) for 30 min. Finally, the gel was dried at 70°C for 2 h (gel dryer 583; Bio-Rad), and the dried gel was subjected to autoradiography.

Turkki P., Laajala M., Flodström-Tullberg M, & Marjomäki V. (2020). Human Enterovirus Group B Viruses Rely on Vimentin Dynamics for Efficient Processing of Viral Nonstructural Proteins. Journal of Virology, 94(2), e01393-19.

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