Fluorescence experiments were performed by utilizing a JASCO FP 8300 spectrofluorometer (JASCO, Tokyo, Japan). Experiments were carried out at 25 °C in 1 cm path-length quartz cuvette. Peptide 4 μM was taken in 30 mM buffer of pH 7.0 containing 100 mM NaCl or 100 mM KCl, 0.5 mM EDTA with and without 40 wt% PEG 200 titrated with equimolar concentration of c-Myc G-quadruplex. The temperature of the cell holder was regulated by a JASCO ETC-273T temperature controller. Samples were prepared by same procedure. Excitation and emission slit width were 5 nm each and the samples were excited at 275 nm and the emission was recorded in a range of 300 nm to 500 nm. Experiment has been repeated in triplicates to reproduce the data. The fluorescence intensity of QW10 was plotted at 342 nm, 347 nm and 343 nm against DNA concentration in the presence of K+ alone or with 40 wt% PEG 200 or 20 wt% PEG 8000.
Etc 273t temperature controller
Manufactured by Jasco
Sourced in Japan
The ETC-273T is a temperature controller designed for laboratory applications. It features a digital display and provides precise temperature control functionality.
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Lab products found in correlation
4 protocols using etc 273t temperature controller
1
Fluorescence-based Characterization of c-Myc G-quadruplex
2
Nanogel Characterization Using TEM and DLS
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TEM images were obtained with a Hitachi H-7100 transmission electron microscope (Hitachi High-Technologies, Tokyo, Japan). A drop of the nanogel solution in ethanol or water (0.01 w/v%, 5 μL) was placed on a formvar-coated copper grid. The specimen was air-dried at room temperature and then examined at an accelerating voltage of 75 kV. The hydrodynamic diameter and zeta potential were measured with a Malvern Instruments Zetasizer Nano ZS (currently Malvern Panalytical, Malvern, UK). The samples were equilibrated for 10 min at each temperature. The amount of fluorescent DBD-AA units in NANOGEL-3 ~6 was estimated from the comparison of the absorbance of their methanol solution with that of N,2-dimethyl-N-(2-{methyl[7-(dimethylsulfamoyl)-2,1,3-benzoxadiazol-4-yl]amino}ethyl)propenamide (ε = 11,000 M−1 cm−1 at 444 nm) as a model compound [33 (link)]. The fluorescence spectra of NANOGEL-3 ~6 were recorded in water and a 150 mM KCl solution at various temperatures using a JASCO FP-6500 spectrofluorometer (Tokyo, Japan) with a Hamamatsu R-7029 optional photomultiplier tube (Hamamatsu, Japan, operating range, 200–850 nm). The sample temperature was controlled using a JASCO ETC-273T temperature controller (Tokyo, Japan).
3
Fluorescence Spectra of Thermometers
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The fluorescence spectra of the synthesized fluorescent thermometers were recorded in KCl solutions at various temperatures using a JASCO FP-6500 spectrofluorometer. The sample temperature was controlled using a JASCO ETC-273 T temperature controller.
4
Intracellular Thermometry Using Fluorescent Probe
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The fluorescence spectra of thermometer 1 in MOLT-4 cells were recorded at various temperatures using a JASCO FP-6500 spectrofluorometer. The cells treated with the thermometer were suspended in PBS in a cuvette (4 mL), and the cell suspension was stirred with a micro round magnetic stir bar (diameter: 2 mm; stirring speed: 800 rpm). The sample temperature was controlled using a JASCO ETC-273 T temperature controller. The temperature resolution (δT ) of 1 was evaluated using the following equation: 28 δT ¼ @T @F ratio δF ratio ð2Þ
where ∂T/∂F ratio and δF ratio represent the inverse of the slope of the fluorescence ratio-temperature diagram and the SD of the fluorescence ratio, respectively. The SD value was obtained by triplicate measurements of one sample at each temperature.
The total cell number was approximately 100, and the cells showing a fluorescence intensity higher than the threshold (maximum autofluorescence intensity) were counted as the cells containing the thermometer. The temperature resolution in intracellular thermometry of HEK293T cells was evaluated using the averaged fluorescence ratios of 1 in a HEK293T cell extract (0.01 w/v%). The HEK293T cell extract was prepared using a previously described procedure. 3
where ∂T/∂F ratio and δF ratio represent the inverse of the slope of the fluorescence ratio-temperature diagram and the SD of the fluorescence ratio, respectively. The SD value was obtained by triplicate measurements of one sample at each temperature.
The total cell number was approximately 100, and the cells showing a fluorescence intensity higher than the threshold (maximum autofluorescence intensity) were counted as the cells containing the thermometer. The temperature resolution in intracellular thermometry of HEK293T cells was evaluated using the averaged fluorescence ratios of 1 in a HEK293T cell extract (0.01 w/v%). The HEK293T cell extract was prepared using a previously described procedure. 3
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