Spectrofluorometer

The sensitivity of the His-AuNCs-based colloidal sensor towards Fe²⁺ was evaluated by incubating for 15 min the colloidal His-AuNCs (350 μL at initial synthesis concentration) with a solution containing Fe²⁺ at concentration ranging between 0 and 44 mM. Afterwards, the PL intensity of the His-AuNCs in the presence of different concentrations of Fe²⁺ was analyzed via a JASCO spectrofluorometer under 380 nm excitation. The degree of PL quenching was correlated with the Fe²⁺ concentration in order to obtain the calibration curve. The LOD was calculated using the following formula [41 (link)]: $$LOD=3.3\frac{S_y}{S}$$
where Sy is the standard deviation of the response and S represents the slope of the obtained calibration curve. The experiments were repeated three times.

The properties of bioactive substances in wines were determined by using three-dimensional (3D-FL) fluorescence (model FP-6500, Jasco spectrofluorometer, serial N261332, Tokyo, Japan). The 3D-FL was measured at emission wavelengths between 200 and 795 nm, and the initial excitation wavelength was 200 nm. For comparison of the obtained results, catechin and gallic acid were used [71 (link)]. Standard phenolic solutions, such as gallic acid and catechin, were prepared daily by dissolving at a concentration of 10 mM in methanol and then diluting with 10 mM phosphate buffer at pH 7.4. The initial fluorescence intensities of fibrinogen, albumin and globulin were measured before their interactions with the investigated wines. The decreases in the fluorescence intensities were used in the estimation of the binding activities.

Fluorometric measurements were used for the evaluation of binding properties of dragon fruit extracts to human serum albumin (HSA). Two (2D-FL) and three-dimensional (3D-FL) fluorescence measurements for dragon fruits extracts at a concentration of 0.17 mg/mL were recorded on a model FP-6500, Jasco spectrofluorometer, Tokyo, Japan, equipped with 1.0 cm quartz cells and a thermostatic bath. The 2D-FL measurements were taken at emission wavelengths from 310 to 500 nm and at excitation of 295 nm. The 3D-FL spectra were collected with subsequent scanning emission spectra from 200 to 550 nm at 1.0 nm increments by varying the excitation wavelength from 200 to 500 nm at 10 nm increments. Gallic acid and epicatechin were used as standards [35 (link)]. 2.0 × 10⁻⁵ mol/L HSA; 0.05 mol/L Tris–HCl buffer with 0.1 mol/L NaCl (pH 7.4) were used for the reaction. The binding properties (%) for fluorescence intensity of peaks a and b were calculated by the comparison of the fluorescence intensities between initial HSA and the final one after interaction with the investigated dragon fruits extracts.

Colony forming units (CFU) of UPEC were measured using recombinant UPEC strain expressing EGFP. The EGFP expressing recombinant UPEC strain was created by electroporation of pluxI-EGFP plasmids (streptomycin resistance, sm^r). Briefly, electrocompetent cells were prepared by washing cells growing in log phase with 10% glycerol at 4 °C as previously described52 (link). The plasmid was electroporated using a GenePulserXcell^TM (BIORAD, USA) at 2000 V with a gap between electrodes of 1 mm, 200 Ω resistance, and 25 μF capacitance. The electroporated cells were allowed to recover in LB broth at 37 °C for 1 h. The recovered cells were plated on 1.5% LB-agar plates supplemented with streptomycin and incubated at 37 °C overnight. The EGFP fluorescence of cultures was then measured by using a spectrofluorometer (JASCO, Japan) or microtiter plate reader (BioTek, USA).

Photoluminescence measurements were performed using JASCO 6000 Spectro-fluorometer utilizing 150W Xe-tube UV light source. Radio-luminescence was measured by replacing UV light by ⁹⁰Sr-⁹⁰Y (7.74 kBq) beta radiation source as excitation source. Transmittance was investigated by using Analytical Jena Specord-900 photodiode array spectrophotometer.