Fluorolog 3 11

Transmission electron microscopy (TEM) observations were performed on a Tecnai F20 microscope. Atomic force microscope (AFM) measurements were carried out with Veeco Dimension 3100V. X-ray photoelectron spectroscopy (XPS) was carried out with ESCALAB 250Xi (Thermo Scientific, Waltham, MA, USA). Fourier transform infrared (FT-IR) spectra were obtained on a Nicolet 6700 FT-IR spectrometer (Thermo Nicolet Corp., Madison, WI, USA). Photoluminescence emission and excitation spectra were measured on a Hitachi F-4600 spectrophotometer (Hitachi, Tokyo, Japan) equipped with a Xe lamp at ambient conditions. UV-Vis absorption spectra were recorded on a PERSEE T10CS UV-Vis spectrophotometer (Persee, Beijing, China). PL lifetime was measured using Fluorolog 3-11 (HORIBA Jobin Yvon, Kyoto, Japan). PL Quantum yields were measured on a QE-2100 quantum efficiency measurement system (Otsuka Electronics, Japan). Photographs were taken using a Canon camera (EOS 550, Tokyo, Japan) under excitation by a hand-held UV lamp (365 nm).

IR spectra of co-oligomers were taken on FT-IR spectrophotometer (Shimadzu, Model IRA Affinity-1) in the form of KBr pellets. The integrated absorption coefficient was determined using the IRA Affinity-1 software through Gaussian Lorentzian curve fittings. Ultraviolet-visible light (UV-vis) spectra were taken on UV-vis spectrophotometer (Shimadzu, Model UV-1800). The viscosity of the co-oligomers was determined at 25 °C temperature using Ubbehlode viscometer. XRD patterns of the co-oligomers were recorded on a powder diffractometer (Philips, Model PW 3710) (using a nickel-filtered Cu-Kα radiation). Peak parameters were analyzed via Origin Pro 8 software. Fluorescence studies were performed on fluorescence spectrophotometer Fluorolog @ 3–11 (Horiba) The quantum yield was calculated as per the method reported in our earlier studies.^{4 (link)}

Fluorescence spectra were measured on Fluorolog 3.11 (Horiba) for tryptophan (NATA) at 22 μM in water, 1:1 water:methanol, methanol, propan-2-ol, while for SD at 2 μM and eIF4E at 2.75 μM in buffer, in a thermostatted micro-cuvette (Hellma) at 15 °C, with excitation wavelength of 280 nm. Concentrations of proteins and NATA were chosen to have equimolar amounts of tryptophan residues in each sample. Energy at maximum of the fluorescence spectra was calculated as: $$E=h\cdot c/{\lambda }_{\mathit{max}},$$ where h was the Planck constant, c was the speed of light in vacuum, λ_max was the wavelength of the maximum.
During the tryptophan-based thermofluorescence assay, the fluorescence emission signals at 350 and 330 nm were measured every 10 s with the integration time of 4.6 s, at the excitation wavelength of 280 nm. The temperature gradient was 1 °C/min. The actual temperature was measured inside the cuvette with the thermocouple and recorded by the thermostat. The assay for SD was repeated five times at different SD concentrations, from 1 to ~13 μM. The assay for eIF4E was performed at 2.75 μM. The melting temperature was determined from a non-linear fitting of the Boltzmann sigmoidal curve: $$F_{350}/F_{330}\left(T\right)={\mathit{FF}}_i+\left({\mathit{FF}}_f-{\mathit{FF}}_i\right)/\left(1+exp\left(\left(T_m-T\right)/\mathit{\text{slope}}\right)\right),$$ where T_m was the melting temperature in °C, slope was a cooperativity coefficient, FF_i and FF_f were the initial and final plateau, respectively.