Fp 8500

Manufactured by Jasco

Sourced in Japan, United States, Germany

The FP-8500 is a fluorescence spectrometer designed for accurate and reliable measurement of fluorescence properties. It provides high-performance fluorescence detection capabilities for a wide range of applications.

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Close spelling variants of this product

FP-8500 spectrofluorometer (89 citations) FP-8500 fluorescence spectrometer (15 citations) FP-8500 spectrofluorimeter (10 citations) FP-8500 fluorometer (7 citations) FP-8500 spectrometer (6 citations) FP-8500 fluorimeter (6 citations) FP-8500 spectrophotometer (4 citations) Model FP-8500 (3 citations) FP-8500 fluorescence spectrophotometer (3 citations) FP-8500 Fluorospectrometer (2 citations)

117 protocols using fp 8500

In vitro aggregation assays of GAPDH and OmpA

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Aggregation of denatured GAPDH from rabbit muscle (Sigma; G-2267) was measured as described previously (Saio et al., 2014 (link)). 125 µM GAPDH was denatured by 3 M guanidine-HCl in 20 mM potassium phosphate, pH 7.0, 100 mM KCl, 4 mM β-mercaptoethanol, 0.5 mM EDTA, and 0.05% NaN₃ for 12 hr at 4°C. The denatured GAPDH was diluted 50-fold into the buffer that does not contain guanidine-HCl and aggregation was monitored by 90° light scattering at 620 nm on a spectrofluorometer (FP-8500, JASCO Corporation) in the absence or presence of TF or TF^mon at the concentration of 0.5 µM or 1 µM. The experiment was carried out at 20°C. The reproducibility was confirmed by independent assays repeated three times.
In anti-aggregation assay on OmpA^1-192, 62 µM OmpA^1-192 in 50 mM Tris-HCl pH 8.0, 500 mM NaCl, 400 mM imidazole, and 8 M urea was diluted 20-fold into 20 mM potassium phosphate, pH 7.0, 100 mM KCl, 4 mM β-mercaptoethanol, 0.5 mM EDTA, and 0.05% NaN₃. Aggregation was monitored by 90° light scattering at 620 nm on a spectrofluorometer (FP-8500, JASCO Corporation) in the absence or presence of TF or TF^mon at the concentration of 4 µM. The experiment was carried out at 25°C.

Saio T., Kawagoe S., Ishimori K, & Kalodimos C.G. (2018). Oligomerization of a molecular chaperone modulates its activity. eLife, 7, e35731.

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OM Permeability Assessment of Ursolic Acid

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The ability of ursolic acid to permeabilize outer membrane of Enterobacteriaceae was assessed by 1-N-phenylethylamine (NPN) uptake assay as reported earlier (Helander and Mattila-Sandholm 2000 (link)). NPN exhibits enhanced fluorescence in phospholipid environment. Since the outer membrane (OM) of Gram negative bacteria affords steric hindrance to hydrophobic molecules and prevents NPN entry due to LPS, increased NPN fluorescence due to treatment, indicates enhanced OM permeability. Briefly, cells were grown to mid-log phase collected and washed with 5 mM HEPES buffer containing 0.2% glucose at pH 7.5 and resuspended in an equal volume of the same buffer. NPN was added at a concentration of 0.5 mM, this was immediately followed by addition of ursolic acid. Fluorescence due to NPN was measured (Ex 350 and Em 420 nm) using spectrofluorimeter (JASCO FP-8500, Jasco, Tokyo, Japan). NPN in buffer and NPN in buffer along with cells were maintained as controls.

Sundaramoorthy N.S., Mohan H.M., Subramaniam S., Raman T., Selva Ganesan S., Sivasubamanian A, & Nagarajan S. (2019). Ursolic acid inhibits colistin efflux and curtails colistin resistant Enterobacteriaceae. AMB Express, 9, 27.

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Fluorescence Titration of ClpB NBD1-M Variants

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Fluorescence titrations were performed at 25 °C in buffer A using a JASCO FP-8500 fluorescence spectrometer (JASCO Germany GmbH) as described previously (25 (link)). The excitation wavelength was set to 296 nm to facilitate selective excitation of protein-bound MANT-dADP via FRET from nearby tryptophan residues. The MANT fluorescence signal was monitored at 441 nm. Direct titrations of ClpB NBD1-M variants (at 2 or 20 μm) with MANT-dADP (2–50 μm) were used to determine the binding affinity of MANT-dADP, which was subsequently applied as the reference K_D in displacement titrations to determine K_D(ADP) or K_D(ATP). Here, ClpB NBD1-M variants (at 2 or 20 μm) were incubated with MANT-dADP (15–40 μm) and subsequently titrated with ADP (2.5–300 μm) or ATP (125–20,000 μm). ATP titrations were performed in the presence of 2 mm phosphoenolpyruvate and 0.01 mg/ml pyruvate kinase (Roche Applied Science) as an ATP-regenerating system. The data were corrected for dilution effects and analyzed with a cubic equation for competing ligands using the initial concentrations of protein and MANT-dADP as well as the K_D(MANT-dADP) from the direct titration as input values (31 (link)). The program GraFit version 5.0 was used for data fitting.

Zeymer C., Fischer S, & Reinstein J. (2014). trans-Acting Arginine Residues in the AAA+ Chaperone ClpB Allosterically Regulate the Activity through Inter- and Intradomain Communication. The Journal of Biological Chemistry, 289(47), 32965-32976.

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Membrane Depolarization by Ursolic Acid and Colistin

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The effect of ursolic acid alone/with colistin in perturbing membrane potential was evaluated using DiSc3, a cationic membrane permeabilizing dye. Intact bacterial cells accumulate the dye in the lipid bilayer, resulting in quenching of fluorescence. When the membrane gets depolarized, dye gets released to the surrounding aqueous phase and fluorescence gets enhanced (Te Winkel et al. 2016 (link)). The fluorescent intensity (Ex 610 ± 5 nm and Em 660 ± 5 nm) of buffer with DiSc3 (1 μM) was measured initially. Mid log cells were added, which reduces the fluorescent intensity due to accumulation of dye in cells. Colistin, ursolic acid and colistin with ursolic acid treatments were given and the resulting variation in fluorescence intensity due to various treatments was quantified using spectrofluorimeter (JASCO FP-8500, Jasco, Tokyo, Japan).

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Spectrofluorometric Analysis of BSA-Au Conjugates

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The luminescence of the BSA–Au conjugates was measured using a Jasco FP8500 spectrofluorometer (Jasco International Co., Ltd., Tokyo, Japan) at 25 °C in MQ water and 360 ± 5 nm excitation in the 380–750 nm range. The spectral correction function was assessed by the Maroncelly dye setup [15 (link)]. The luminescence quantum yields were determined relative to the 0.546 value of quinine sulfate in 1N sulfuric acid [16 (link)]. It must be noted that considering the low sensitivity of the fluorometer in the red region, the red band’s maxima, and the corresponding fluorescence yields, may be slightly underestimated. The air-saturated samples were measured in a 3 mm × 3 mm × 40 mm quartz cuvette with an optical density at the excitation wavelength around 1.4.

Fehér B., Mihály J., Demeter A., Almásy L., Wacha A., Varga Z., Varga I., Pedersen J.S, & Bóta A. (2022). Advancement of Fluorescent and Structural Properties of Bovine Serum Albumin-Gold Bioconjugates in Normal and Heavy Water with pH Conditioning and Ageing. Nanomaterials, 12(3), 390.

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Fluorescence Polarization Assay for Hormone Binding Kinetics

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Hormone binding kinetics were monitored by fluorescence polarization on a Jasco FP-8500 fluorescence spectrometer (Jasco, Groß-Umstadt, Germany) equipped with polarizers. 1 µM apo-GRLBDm, after extensive dialysis to remove dexamethasone as described, was added to various chaperone mixtures with a chaperone and cofactor concentration of 3 µM^{35 (link),38 (link)}. Binding kinetics to 50 nM fluorescently labelled dexamethasone (F-DEX, Thermo Fischer Scientific, Bremen, Germany) were recorded at 20 °C in 20 mM HEPES, 20 mM KCl, 5 mM MgCl₂, 2 mM ATP, pH 7.5. Hormone binding rates were determined by fitting association kinetics to exponential models and the error bars represent the standard deviation of three independent measurements.

Kaziales A., Barkovits K., Marcus K, & Richter K. (2020). Glucocorticoid receptor complexes form cooperatively with the Hsp90 co-chaperones Pp5 and FKBPs. Scientific Reports, 10, 10733.

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Synthesis and Luminescent Properties of Eu-Doped BaSiO3 Phosphors

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A series of Ba₀.₉₈SiO₃:0.02Eu, Ba₀.₉₆SiO₃:0.02Eu, and 0.02R (R = Li, K, Na, La, and Y) phosphors were prepared using a conventional solid-state reaction. The BaCO₃ (99+%), SiO₂ (particle size 5~20 nm, 99.5%), Eu₂O₃ (99.99%), K₂CO₃ (99+%), Li₂CO₃ (99+%), Li₂O (97%), Na₂CO₃ (99+%), LaCl₃ (99.99%), and Y₂O₃ (99.99%), purchased from Merck, were used as a raw materials but SiO₂ was used 20% more to obtain a single-phase BaSiO₃ sample. The raw materials were thoroughly mixed in an agate mortar and subsequently fired in air at 1100 °C for 4 h to synthesize the Eu³⁺-doped BaSiO₃. The BaSiO₃:Eu²⁺ samples were prepared by sintering the BaSiO₃:Eu³⁺ samples again under a reduction atmosphere (95% N₂ + 5% H₂) at 1200 °C for 6 h. The specific formulations of the studied samples are shown in Table 1.
The crystal phase of the obtained samples was identified using an X-ray diffractometer (X’Pert PRO MPD-3040, Malvern Panalytical, Malvern, UK, CuK_α1, λ = 1.5406 Å) operating at 40 kV and 30 mA with a scan speed of 0.02°/sec. The photoluminescence excitation (PLE) and emission (PL) spectra were measured using a fluorescent spectrofluorometer (JASCO FP-8500, JASCO, Tokyo, Japan) equipped with an integrating sphere (ISF-834, JASCO, Tokyo, Japan). All measurements were performed at room temperature.

Namkhai P, & Jang K. (2022). Co-Doping Effect on the Optical Properties of Eu(2+/3+) Doped in BaSiO3. Materials, 15(19), 6559.

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Protein Folding State Verification

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The folded state of proteins and
their conjugates was verified by spectrofluorimetry. The fluorescence
spectra were acquired using an FP-8500 spectrofluorimeter (Jasco,
Japan) with excitation at 280 nm and emission in the 300–450
nm range, at a protein concentration of ∼4 × 10^–6 M in Dulbecco’s PBS.

Krzyscik M.A., Zakrzewska M., Sørensen V., Sokolowska-Wedzina A., Lobocki M., Swiderska K.W., Krowarsch D., Wiedlocha A, & Otlewski J. (2017). Cytotoxic Conjugates of Fibroblast Growth Factor 2 (FGF2) with Monomethyl Auristatin E for Effective Killing of Cells Expressing FGF Receptors. ACS Omega, 2(7), 3792-3805.

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In vivo eGFP Fluorescence Measurements

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The in vivo eGFP fluorescence measurements were performed in E. coli SQ171. Precultures were incubated in 5 ml LB medium with 200 µg ml -1 trimethoprim over night at 37 • C and 200 rpm. 100 µl of the precultures were used to inoculate fresh cultures (5 ml), which were incubated at 37 • C and 200 rpm until an OD 600 of 0.5. Expression of eGFP was induced by addition of L-(+)-arabinose (to 0.1% w/v) in the absence and presence of neomycin (250 µg ml -1 ). After incubation for 3 h at 37 • C and 200 rpm, cultures were immediately stored on ice to measure the final OD 600 and diluted with cold 1× PBS to 1 ml (OD 600 = 1.5). Samples were centrifuged at 8 000 rpm for 5 min, the pellet was washed twice with cold 1× PBS and resuspended in 500 µl 1× PBS (equaling OD 600 = 3.0). eGFP fluorescence was measured from 503 nm to 600 nm with a fluorescence spectrometer (JASCO Deutschland GmbH, model: FP-8500, mode: emission, ex bandwidth: 2.5 nm, em bandwidth: 5 nm, response: 1 s, sensitivity: low, ex wavelength: 488 nm).
As a positive control, we combined a 5' adenosine residue following the transcription start site (TSS) with the poly-U stretch of length eight and the terminator downstream region (TDR) of our riboswitch constructs. The poly-U stretch is known to promote translation [16] (link) and, thus, makes the control comparable to the full riboswitch sequences.

Günzel C., Kühnl F., Arnold K., Findeiß S., Lünse C.E., Stadler P.F., & Mörl M. (2020). Beyond Plug and Pray: Context Sensitivity and in silico Design of Artificial Neomycin Riboswitches.

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Tryptophan Fluorescence Quenching Assay for Protein-Ligand Binding

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The tryptophan fluorescence quenching assay can be used to assess small molecule inhibitor binding affinities to proteins using fluorescence spectroscopy [65 (link)].
Tryptophan fluorescence quenching assays were performed in black 96-well plates. The CLuc-NLRP3^PYD chimeric protein was added to each well, and then different concentrations (2, 10, 20, and 50 µM) of QM381 compound were loaded in the wells. The fluorescent signal of each well was detected at the same time. NLRP3^PYD excitation at 280 nm prompts an emission maximum corresponding to the tryptophan fluorescence at 352 nm in a Jasco FP-8500 spectrofluorometer.

Moasses Ghafary S., Soriano-Teruel P.M., Lotfollahzadeh S., Sancho M., Serrano-Candelas E., Karami F., Barigye S.J., Fernández-Pérez I., Gozalbes R., Nikkhah M., Orzáez M, & Hosseinkhani S. (2022). Identification of NLRP3PYD Homo-Oligomerization Inhibitors with Anti-Inflammatory Activity. International Journal of Molecular Sciences, 23(3), 1651.

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