ABSTRACT
BACKGROUND: Golgi-Cox staining has been conventionally used for investigating neuronal development. After the brain tissue is subject to Golgi-Cox staining, black deposits are formed on the surface of the stained neurons because of mercuric sulfide, which does not show a fluorescence response under two-photon excitation. However, we unexpectedly observed fluorescence emitted by these black deposits during two-photon fluorescence measurements. Further, the in-depth of physical and chemical methods analysis revealed that the black deposits on the stained neurons are composed of Hg-binding proteins. METHODS: We studied black deposits present in the Golgi-Cox-stained mouse brain neurons using techniques such as multiple-photon microscopy, scan electron microscopy, micro-Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. RESULTS: The emitted fluorescence was because of the fluorescence groups of Hg-binding protein present within the Golgi-Cox deposits on the neuronal surface. CONCLUSIONS: The presence of Hg-binding proteins within black deposits on the surface of Golgi-Cox-stained neurons was proven for the first time. The novel interaction between the neurons and Hg2+ ions during Golgi-Cox staining help to understand the mechanism of Golgi-Cox staining.
Subject(s)
Brain/metabolism , Carrier Proteins/metabolism , Golgi Apparatus/metabolism , Mercury/metabolism , Neurons/metabolism , Staining and Labeling/methods , Animals , Brain Chemistry/physiology , Carrier Proteins/analysis , Female , Golgi Apparatus/chemistry , Male , Mercury/analysis , Mice , Mice, Inbred C57BL , Microscopy, Fluorescence, Multiphoton/methods , Neurons/chemistry , Photoelectron Spectroscopy/methodsABSTRACT
The unstable-waveguide hybrid resonator emits a rectangular, simple astigmatic beam with a large number of high-spatial-frequency oscillations in the unstable direction. To equalize the beam quality, in this paper, a beam shaping system with a spatial filter for the hybrid resonator was investigated by numerical simulation and experimental method. The high-frequency components and fundamental mode of the output beam of the hybrid resonator in the unstable direction are separated by a focus lens. The high-frequency components of the beam are eliminated by the following spatial filter. A nearly Gaussian-shaped beam with approximately equal beam propagation factor M² in the two orthogonal directions was obtained. The effects of the width of the spatial filter on the beam quality, power loss, and intensity distribution of the shaped beam were investigated. The M² factor in the unstable direction is changed from 1.6 to 1.1 by optimum design. The power loss is only 9.5%. The simulation results are in good agreement with the experimental results.
