Fluorescence spectra were acquired using a Jasco-FP8300 spectrofluorometer (Jasco Inc., Easton, MD, USA) equipped with a temperature-controlled circulator. Fluorescence was measured in a quartz cell with path length of 1 cm, using an excitation wavelength of 377 nm and an emission spectra scan from 520–600 nm. The titration experiments were carried out at protoberberine concentrations of 0.2 μM in 100 mM K+. wtTel26 DNA at the specified concentrations was titrated into the four protoberberine solutions, respectively, and the resulting solutions were incubated for 2 min before fluorescence measurements were taken. The disassociation constant Kd was calculated using GraphPad Prism software (San Diego, CA, USA fitting of an equation: A = Amin + (Amax − Amin) [(PT + OT + Kd) − [((PT + OT + Kd)2 − (4PTOT))1/2]/(2OT), where A represents the fluorescence intensity of the protoberberines bound to wtTel26 DNA. Both Amax and Amin are fit in addition to Kd. The protoberberine concentration, OT, was held constant, and PT, the total complex concentration is the independent variable, varying with each measurement of A.
Fp 8300 spectrofluorometer
Manufactured by Jasco
Sourced in Japan, United States, Germany, Italy
The FP-8300 spectrofluorometer is a laboratory instrument designed to measure the fluorescent properties of samples. It is capable of recording both excitation and emission spectra of fluorescent materials.
Automatically generated - may contain errors
Lab products found in correlation
Pct 818, by Jasco (5 mentions)
V 750 spectrophotometer, by Jasco (3 mentions)
Avance 300, by Bruker (3 mentions)
Spectra manager software, by Jasco (2 mentions)
B 540 apparatus, by Büchi (2 mentions)
Avance 500, by Bruker (2 mentions)
Uv 2501pc spectrophotometer, by Shimadzu (2 mentions)
Lambda 650, by PerkinElmer (2 mentions)
Ctu 100, by Jasco (2 mentions)
Pac 743r, by Jasco (2 mentions)
Zuv h30m head unit, by Omron (2 mentions)
Prism software, by GraphPad (2 mentions)
Borate buffer, by Merck Group (1 mentions)
Etc 273t, by Jasco (1 mentions)
Calcium calibration buffer kit, by Thermo Fisher Scientific (1 mentions)
Avance 3 300 mhz spectrometer, by Bruker (1 mentions)
Ft ir 4700 spectrometer, by Jasco (1 mentions)
Etc 273t temperature controller, by Jasco (1 mentions)
Uv 2450 uv vis spectrophotometer, by Shimadzu (1 mentions)
Fs5 fluorescence spectrophotometer, by Edinburgh Instruments (1 mentions)
115 protocols using fp 8300 spectrofluorometer
1
Protoberberine Binding Affinity to G-Quadruplex DNA
2
Peptide Aggregation Propensity Evaluation
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To study the aggregation propensity of the peptides in the absence and in the presence of the metal complexes, thioflavin (ThT) fluorescence time-course experiments were carried out at 25 °C using a Jasco FP 8300 spectrofluorometer (JASCO, Tokyo, Japan). λexc: was set to 440 nm, while λem was set to 483 nm. ThT concentration was 50 µM, peptide concentration was 100 µM. The solution was maintained under magnetic stirring in a 10 mm path-length quartz cuvette. Experimental conditions: 0.1% DMSO, 10 mM borate buffer (Merck KGaA, Darmstadt, Germany) at pH = 9.0 for NPM1264–277 and 0.1% DMSO, 10 mM sodium phosphate buffer for Aβ21–40. Reported fluorescence values are the mean of two independent experiments.
3
Characterizing Myc2345 DNA Binding
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All fluorescence spectra were acquired on a Jasco FP-8300 spectrofluorometer (JASCO, Inc.) equipped with a temperature controller at 25°C. A 100 nM sample of 3′-TAMRA-labeled Myc2345_T23 was prepared in 25 mM potassium phosphate buffer, 75 mM KCl, pH 7. The emission spectrum between 565 and 610 nm was recorded with an excitation wavelength of 555 nm, excitation and emission bandwidth of 5 nm, a response time of 4 s, a data interval of 1 nm, and a scanning speed of 50 nm/min. An initial titration of PEQ to the 3′-TAMRA-labeled Myc2345_T23 was performed to determine the optimal ligand-DNA ratio for the competition experiments. A 10:1 ratio was chosen to obtain significant TAMRA quenching while limiting the excess of PEQ in solution. The emission at 579 nm was used to follow the changes in TAMRA fluorescence upon ligand or DNA addition. Myc2345 or Myc2345_T23 were titrated to the 10:1 mixture of PEQ and 3′-TAMRA labeled Myc2345_T23 in duplicate to determine the C50 value, the concentration of the competing unlabeled DNA sequence at which half the initially quenched fluorescence is restored. For this purpose, the normalized change in TAMRA fluorescence ΔF was fitted against ΔFmax/(1 + (C50/x)n) with ΔFmax as maximum change in fluorescence, x as the concentration of the competing DNA sequence, and n as slope.
4
Fluorescence Quenching Analysis of Peptide-Ligand Interaction
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Fluorescence experiments were performed by utilizing a JASCO FP 8300 spectrofluorometer (JASCO, Tokyo, Japan). Experiments were carried out at 25 °C in a 3 mm path-length quartz cuvette for 4 μM peptide in pH 7.0 buffer containing 100 mM KCl, 0.5 mM EDTA titrated with equimolar concentration of HTPu. 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. Modified Stern–Volmer equation was used to analyze fluorescence quenching data to find out various binding parameters for this interaction since binding parameters are vital to study about the binding mechanism.where the highest fluorescence intensity in the absence of ligand is F0 whilst F depicts fluorescence intensity in the presence of ligand, K depicts the binding constant, n depicts the number of binding sites, and the concentration of RT is depicted by C.
5
Fluorescence Quenching Assay for Berberine Binding
6
Spectroscopic Measurements in Quartz Cuvette
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All spectroscopic measurements were carried out in a 700 μl quartz cuvette at room temperature. Absorption spectra were recorded with the JASCO V-750 spectrophotometer (JASCO GmbH). Emission spectra were recorded with the Jasco FP-8300 spectrofluorometer (JASCO GmbH). To measure spectra in the dark samples were handled under a red safety light.
7
Fluorescence Titration Assay of BMVC Binding
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Fluorescence spectra were acquired with a Jasco-FP8300 spectrofluorometer (Jasco Inc., Easton, MD, USA) equipped with a temperature-controlled circulator. Fluorescence was measured between 500 and 600 nm in a quartz cell with path length of 1 cm, using an excitation wavelength of 475 nm. The titration experiments were carried out at a BMVC concentration of 20 nM in presence of 25 mM K-phosphate and 70 mM KCl buffer or 25 mM Na-phosphate and 70 mM NaCl buffer at pH 7. Six different DNA sequences at the specific concentration were added into the BMVC solutions and the resulting solutions were incubated for 2 min before fluorescence was measured. The apparent Kd was calculated using GraphPad Prism software to fit the equation assuming a 1:1 model: F = Fmax [(MT + LT + Kd) – ((MT + LT + Kd)2 – (4MTLT))1/2]/(2LT), where F represents the BMVC fluorescence intensity at 563 nm. Fmax is fitted in addition to apparent Kd. The total BMVC (ligand) concentration, LT, was held constant and MT, the total DNA (Macromolecule) concentration is an independent variable, varying with each titration step. The binding isotherm could be fitted with the equation assuming a 1:1 model, suggesting that the apparent Kd is probably dominated by the stronger binding site at the 5′-end.
8
Spectrofluorometric Characterization of ZnO NCs
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The ZnO NCs were analyzed with a JASCO FP-8300 spectrofluorometer (JASCO Europe, Cremella, Italy) and the three-dimensional (3D) excitations and emission spectra were recorded with 5 nm wavelength interval in the 210–735 nm range. As a control sample, zinc nitrate and water were used.
9
Spectroscopic Characterization of Fluorescent Dyes
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All the solvents were of analytical grade. Chemicals were purchased from commercial sources. 1H-NMR and 13C-NMR were measured on a Bruker Avance III-300 MHz spectrometer (Bruker Biospin, The Woodlands, TX, USA) with chemical shifts reported in ppm (TMS as internal standard). Mass spectra were measured on a Focus GC/DSQ II spectrometer (ThermoScientific, Waltham, MA, USA) for IC and an API 3000 spectrometer (Applied Biosystems, PE Sciex) for ES. All pH measurements were made with a Mettler Toledo pH meter. Fluorescence spectra were recorded on a JASCO FP-8300 spectrofluorometer (JASCO, Easton, MD, USA). Absorption spectra were determined on a VARIAN CARY 300 Bio UV-Visible spectrophotometer. All measurements were done at a temperature of 25°C. The purity of the dyes were checked by RP-HPLC C-18, elutant: ACN 0.1% TFA/Water 0.1% TFA, method: 20/80 to 100/0 within 20 min then 100/0 for 10 min detection at λAbs = 254 nm. The apparent dissociation constant for calcium (KD Ca2+) was measured with a calcium calibration buffer kit from Invitrogen (Lifetechnologies, USA). All mass spectra, NMR spectra and chromatograms are included as supplemental data.
10
Comprehensive Analytical Characterization of Organic Compounds
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Melting point was recorded using Sturat SMP30 (UK). The elemental analysis was measured using EA 1108 FISONS elemental analyzer (Germany). Nuclear Magnetic Resonance (NMR) spectra were recorded on a Bruker-spectroscopy DTX 400 ultrashield in hexadeuterateddimethylsulphoxide (DMSO-d6) using tetramethylsilane (TMS) as internal reference and the chemical shifts (δ) are given in. The Uv/vis absorption spectra were recorded on a shimadzuUV2401 spectrophotometer at the wavelength of maximum absorption (λmax) in a range of solvent with different polarity. Mass spectra were measured using Thermo-Scientific ™ Fleet LCQ ™ ion trap (Germany). Samples were dissolved in MeCN and analyzed by direct infusion at a flow rate of 5 mL/min. Mass spectra were recorded in the range of 50–1000 m/z using electrospray ionization (ESI) and the negative ions as well as positive ion were recorded. Helium 0.5 mL/min was used as a carrier gas. Pressure and flow rate of nitrogen was 10 psi and 5 L/min. Ion source temperature was 275 °C, an ionization voltage was from 2.6 to 2.7 kV.
FT/IR spectra were measured on JASCO FT/IR-4700 spectrometer. Steady-state fluorescence spectra were measured using a JASCO FP-8300 spectrofluorometer (JASCO Inc., USA), the emission spectra was measured in different environment such as polycrystalline, in solution and on the dyed fabric.
FT/IR spectra were measured on JASCO FT/IR-4700 spectrometer. Steady-state fluorescence spectra were measured using a JASCO FP-8300 spectrofluorometer (JASCO Inc., USA), the emission spectra was measured in different environment such as polycrystalline, in solution and on the dyed fabric.
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