Subcellular localization of boron in cultured melanoma cells by electron energy‐loss spectroscopy of freeze‐dried cryosections

Abstract

SummaryBoron neutron capture therapy (BNCT) is based on the ability of the non-radioactive isotope 10 B to capture thermal neutrons and to disintegrate instantaneously. This reaction opens a way to selectively destroy tumour cells after specific uptake of 10 B. In this paper, a method based on electron energy-loss spectroscopy is presented for detecting and quantifying boron in freeze-dried cryosections of human melanoma cells. A practical detection limit of around 6 mmol kg − 1 in 0.1-µ m 2 areas is estimated using specimens prepared from standard boron solutions. Preliminary results of boron mapping in the spectrum-imaging acquisition mode reveal boron penetration and probably spot-like accumulation within melanoma cells when exposed to culture medium containing sodium borocaptate.

Boron analysis and boron imaging in biological materials for Boron Neutron Capture Therapy (BNCT)

“…the best spatial resolution is obtained using methods based on electron microscopy. Two methods exist based on electron microscopy for boron imaging: immunohistochemistry [89] and Electron Energy Loss Spectroscopy [90,91]. Immunochemistry is a very sensitive method but it has its drawbacks: indirect detection leading to the possibility of artifacts (e.g., redistribution during the fixation process) and the need to prepare a specific antibody for each potential boron carrier molecule.…”

“…Secondly, it can be observed that the boron K edge located at 188 eV, can be confused with the phosphorus L1 signal. Consequently, the experimental filtered spectrum has to be modeled as a sum of a boron-filtered reference and a phosphorus-filtered reference spectrum [91]. Neglecting the phosphorus signal can lead to erroneous images, where boron is systematically detected in phosphorus rich regions.…”

“…Taking into account these methodological developments and in our current experimental conditions, the detection limits can be estimated to be typically around a few tens of ppm in, for a sample area of 0.1 m 2 (see ref. [91] for details of the method for boron detection and quantification).…”

Laser postionization secondary neutral mass spectrometry in tissue: a powerful tool for elemental and molecular imaging in the development of targeted drugs

“…Besides minimizing the time between tissue excision and sample freezing, it is important to maximize the cooling rate during the freezing process. A dedicated cryogenic sample preparation technique with a high cooling rate was validated for tissue samples (20,21). Liquid propane was cooled below its boiling temperature using liquid nitrogen, thus accelerating the cooling process and preventing evaporation of propane at the contact surface of the immersed specimen.…”

SIMS ion microscopy imaging of boronophenylalanine (BPA) and 13C15N-labeled phenylalanine in human glioblastoma cells: Relevance of subcellular scale observations to BPA-mediated boron neutron capture therapy of cancer

“…Cell culture tumor models are ideally suited for understanding the mechanisms of boron delivery by BNCT agents [10,11]. Techniques capable of providing subcellular scale boron measurements are critically needed in BNCT [12][13][14][15][16]. The subcellular scale measurements require the use of reliable cryogenic sample preparations so that the native distribution of boron in subcellular compartments can be maintained during sampling.…”