Nanoled

Time-resolved fluorescence (TRF) decays were obtained through a home-built-time-correlated single photon counting (TCSPC) apparatus previously described [24 (link)]. The decays were collected with excitation at 282 nm (HoribaJobin-Yvon-IBH nanoLED) and emission wavelength at 338 nm. The fluorescence decays for HSA (10 μM, in PBS) and HSA/TFV (drug concentrations of 8.2, 10, and 26 μM, in PBS) and HSA/TDF (prodrug concentrations of 8.2, 10, and 26 μM in PBS), as well as the instrumental response function (IRF, collected using a Ludox^® dispersion), were obtained using 1024 channels until 2000 counts at the maximum. Deconvolution of the fluorescence decay curves was performed using the modulation function method, as implemented by G. Striker in the SAND software version 1.0, as previously reported in the literature [25 (link)]. The average fluorescence lifetime (τ_average) was determined following Equation (5): $${\tau }_{\mathrm{average}}=\frac{\sum A_i{\tau }_i^2}{\sum A_i{\tau }_i}$$
where τ_i is the fluorescence lifetime and A_i is the pre-exponential factor.

Samples for optical characterizations were prepared by dispersing them into toluene in 1 cm path length quartz cuvettes. UV–Vis absorption spectra were recorded using a Varian Cary 60 UV–visible spectrophotometer. PL characterization was carried out using a Horiba Jobin Yvon Fluorolog-3 using a Hamamastu InP/InGaAs photomultiplier (R5509-7–3). Excitation wavelength that used for the PL measurements was performed at least 20 nm below the emission range. Photoluminescence quantum yields (PLQYs) were calculated by the comparison of the PL intensity between samples and standard dyes, which have well-known PLQYs. HITCI (PLQY = 30% in ethanol) was used for the determination of the PLQY of the Cu–In–S QDs. Rhodamine 6G (PLQY = 91% in ethanol) was employed as the reference to calculate the PLQY of Cu–In–(Zn)–S/ZnS “ice-cream” like nanoheterostructure. In order to minimize the self-absorption of fluorescence, the absorbance of the samples was measured first to be 0.5 at peak excitation value, followed by 1:10 dilution, then the PL spectra of the samples were recorded. PL decay curves were performed using a time correlated single photon counting (TCSPC) spectrometer (Horiba Jobin Yvon) and the semiconductor diode laser (372 nm “Nano LED − 370”—HORIBA jobin Yvon) with pulse duration shorter than 200 ps for excitation.

Fluorescence spectra were recorded on a Fluorolog-3 JobinYvon-Spex spectrofluorometer (model GL3–21, Kyoto, Japan), using a laser diode as the excitation source (NanoLED, 440 nm, pulse width 100 ps) and a TBX-PMT series UV detector (250–850 nm) from Horiba Jobin Yvon. The micropolarity of the copolymer aggregates in aqueous solution at different pHs was evaluated by utilizing pyrene as the fluorescent probe. Pyrene is sensitive to the polarity of the surrounding environment when residing within the polymer aggregates formed in each case. A quantity of pyrene solution (in acetone, 1 mM) was added to each solution at a ratio of 1 µL/1 mL. The samples were allowed to equilibrate for 24 h (in this period, acetone evaporation takes place) and then measured at ambient temperature.