Simultaneous Observation of Cells and Nuclear Tracks from the Boron Neutron Capture Reaction by UV-C Sensitization of Polycarbonate

Abstract

The distribution of boron in tissue samples coming from boron neutron capture therapy protocols can be determined through the analysis of its autoradiography image on a nuclear track detector. A more precise knowledge of boron atom location on the microscopic scale can be attained by the observation of nuclear tracks superimposed on the sample image on the detector. A method to produce an "imprint" of cells cultivated on a polycarbonate detector was developed, based on the photodegradation properties of UV-C radiation on this material. Optimal conditions to generate an appropriate monolayer of Mel-J cells incubated with boronophenylalanine were found. The best images of both cells and nuclear tracks were obtained for a neutron fluence of 1013 cm-2, 6 h UV-C (254 nm) exposure, and 4 min etching time with a KOH solution. The imprint morphology was analyzed by both light and scanning electron microscopy. Similar samples, exposed to UV-A (360 nm) revealed no cellular imprinting. Etch pits were present only inside the cell imprints, indicating a preferential boron uptake (about threefold the incubation concentration). Comparative studies of boron absorption in different cell lines and in vitro evaluation of the effect of diverse boron compounds are feasible with this methodology.

Neutron Autoradiography Combined With UV-C Sensitization: Toward the Intracellular Localization of Boron

“…As control samples, a set of Lexan foils were exposed to UV-C radiation without previous hematoxylin staining. As it was determined in our previous work (Portu et al, 2015b), cell structures absorb UV-C radiation at different levels, generating reliefs on the etched detector's surface. In the first column of Figure 2, it is noteworthy that cellular imprints are formed from the UV-C absorption of each cellular structure (focusing on the nucleus and the cytoplasm).…”

“…Then, only 5 min of UV-C exposure is enough to obtain high-quality imprint images when hematoxylin is previously used. This fact represents a considerable reduction of irradiation time, in comparison with the 6 h UV-C exposure applied in previous studies (Portu et al, 2015 b ).…”

“…The described protocol could vary when working with other cell lines. Indeed, in previous works performed with melanoma cell lines, a pretreatment with the culture medium for 24 h was necessary to obtain adequate cell distribution on Lexan (Portu et al, 2015 b ; Rossini et al, 2015).…”

“…Fixed cells on the NTD (with or without staining) were irradiated with UV-C at different exposure times: 2, 5, 15, and 30 min and 1, 2, and 6 h. These last two exposure times were included according to previous experiences (Portu et al, 2015 b ). The UV-C irradiation was performed with a 15 W UV lamp with 254 nm wavelength emission (G15T8, General Electric) (Portu et al, 2016).…”

“…Different experiments were carried out in order to establish cell culture conditions on polycarbonate substrates. Furthermore, optimal conditions were set up to simultaneously visualize cell imprints and nuclear tracks coming from the BNC reaction (Portu et al, 2015 b ). On the other hand, a reduction in the nuclear track density (fading) on sensitized polycarbonate foils by UV-C radiation was reported by our group.…”