A Rapidly Modulated Multifocal Detection Scheme for Parallel Acquisition of Raman Spectra from a 2‑D Focal Array

We report the development of a rapidly modulated multifocal detection scheme that enables full Raman spectra (∼500–2000 cm^–1) from a 2-D focal array to be acquired simultaneously. A spatial light modulator splits a laser beam to generate an <i>m</i> × <i>n</i> multifocal array. Raman signals generated within each focus are projected simultaneously into a spectrometer and imaged onto a TE-cooled CCD camera. A shuttering system using different masks is constructed to collect the superimposed Raman spectra of different multifocal patterns. The individual Raman spectrum from each focus is then retrieved from the superimposed spectra with no crosstalk using a postacquisition data processing algorithm. This system is expected to significantly improve the speed of current Raman-based instruments such as laser tweezers Raman spectroscopy and hyperspectral Raman imaging

Direct Analysis of Water Content and Movement in Single Dormant Bacterial Spores Using Confocal Raman Microspectroscopy and Raman Imaging

Heavy water (D₂O) has a distinct molecular vibration spectrum, and this has been used to analyze the water content, distribution, and movement in single dormant Bacillus cereus spores using confocal Raman microspectroscopy and Raman imaging. These methods have been used to measure the kinetics of D₂O release from spores suspended in H₂O, the spatial distribution of D₂O in spores, and the kinetics of D₂O release from spores during dehydration in air at room temperature. The results obtained were as follows. (1) The Raman spectrum of single D₂O-loaded dormant spores suggests that D₂O in spores is in a relatively weak hydrogen-bonded mode, compared to the strong hydrogen-bonded mode in pure D₂O. (2) The D₂O content of individual spores in a population was somewhat heterogeneous. (3) The spatial distribution of D₂O in single dormant spores is uneven, and is less dense in the central core region. Raman images of different molecular components indicate that the water distribution is somewhat different from those of proteins and Ca-dipicolinic acid. (4) Exchange of spore D₂O with external H₂O took place in less than 1 s. (5) However, release of spore D₂O during air dehydration at room temperature was slow and heterogeneous and took 2–3 h for complete D₂O release

Fast Confocal Raman Imaging Using a 2‑D Multifocal Array for Parallel Hyperspectral Detection

We present the development of a novel confocal hyperspectral Raman microscope capable of imaging at speeds up to 100 times faster than conventional point-scan Raman microscopy under high noise conditions. The microscope utilizes scanning galvomirrors to generate a two-dimensional (2-D) multifocal array at the sample plane, generating Raman signals simultaneously at each focus of the array pattern. The signals are combined into a single beam and delivered through a confocal pinhole before being focused through the slit of a spectrometer. To separate the signals from each row of the array, a synchronized scan mirror placed in front of the spectrometer slit positions the Raman signals onto different pixel rows of the detector. We devised an approach to deconvolve the superimposed signals and retrieve the individual spectra at each focal position within a given row. The galvomirrors were programmed to scan different focal arrays following Hadamard encoding patterns. A key feature of the Hadamard detection is the reconstruction of individual spectra with improved signal-to-noise ratio. Using polystyrene beads as test samples, we demonstrated not only that our system images faster than a conventional point-scan method but that it is especially advantageous under noisy conditions, such as when the CCD detector operates at fast read-out rates and high temperatures. This is the first demonstration of multifocal confocal Raman imaging in which parallel spectral detection is implemented along both axes of the CCD detector chip. We envision this novel 2-D multifocal spectral detection technique can be used to develop faster imaging spontaneous Raman microscopes with lower cost detectors

PQQ Inhibited Osteoclastogenesis Caused by Wear Particles in vivo.

<p>(A) Histologic appearance of particle induced in vivo mouse calvarial osteolysis on d 14.Representative HE (a–d) and TRAP (e–h) stained histologic slices are presented at ×40. High power images of osteoclasts in the boxed regions in (f) and (g) are also shown at ×200 (i–l). UHMWPE induced significant bone resorption (b) and osteoclastogenesis (f) than the PBS group (a, e). This inflammatory reaction was effectively inhibited by PQQ (1 mg/kg) (c, g) and PQQ (10 mg/kg) (d, h) treatment (n = 6). (B) UHMWPE particles induced osteolysis area was much bigger than the control. (C) UHMWPE particles induced formation of more TRAP (+) cell than the control. While PQQ could effectively inhibited wear particles induced osteolysis and osteoclastogenesis (n = 6). Asterisk indicates a statistically significant difference (<i>p<0.05</i>), comparison between the two groups connected by the line.</p

Bone histomorphometry parameters after 14-day treatment with PQQ.

<p>Values are mean ±SD (n = 6).</p>*<p><i>P<0.01</i> different from control group.</p>#<p><i>P<0.05</i> different from control group.</p>∧<p><i>P<0.01</i> different from UHMWPE group.</p><p><a href="mailto:@P" target="_blank">@P</a><0.05 different from UHMWPE group.</p>∇<p><i>P<0.05</i> different from low dose PQQ group.</p

PQQ Inhibited Osteolysis Caused by Wear Particles in vivo.

<p>Micro-CT three-dimensional reconstruction images of bone resorption in mouse calvarium. (A) Sham control group, (B) UHMWPE, (C) UHMWPE+PQQ 1 mg/kg, (D) UHMWPE+PQQ 10 mg/kg. Arrow indicates the low signal bone resorption area.</p

Change of bone parameters of mice skull analyzed by micro-CT.

<p>(A) BMD, bone mineral density, (B) BVF, bone volume fraction, (C) CMT, cortical mean thickness, (D) Ct, cortical area/total area data are presented as mean ± SD (n = 6). Asterisk indicates a statistically significant difference (<i>p<0.05</i>), comparison between the two groups connected by the line.</p

PQQ suppresses the mRNA expression of c-Fos, NFATc1, and TRAP in BMMs treated with RANKL.

<p>(A–C) BMMs were pretreated with or without PQQ (15 µM) for 1 h and with RANKL (100 ng/ml) for the indicated periods. The mRNA expression of the indicated genes was analyzed by real time RT-PCR. Data are presented as mean ± SD; 0: Blank Control; <i>^#p</i><0.05 <i>vs.</i> Blank Control; <i>*p</i><0.05 <i>vs.</i> DMSO+RANKL. (D) PQQ inhibits the expression of c-Fos and NFATc1 induced by RANKL. BMMs were pretreated with or without PQQ (15 µM) for 1 h and were treated with RANKL (100 ng/ml) for the indicated periods. Cells were lysed in the lysis buffer, and lysates were analyzed by Western blotting with antibodies against c-Fos, NFATc1, and actin. The intensities of the protein bands were analyzed and normalized to actin. Similar results were obtained in at least 3 independent experiments.</p

Another typical case for group A.

<p>A 45-years-old patient’s preoperative CT scanning shows destructive segments located at C7/T1 segments with collapse of T1 vertebra (a-b). Preoperative sagittal MRI shows the tuberculosis focus is located higher than the suprasternal notch level (c). One-week postoperative X-ray image shows internal fixation in good position (d). Three years postoperative CT scanning reveals cervicothoracic anterior graft fusion (e-f).</p

Effects of PQQ on LPS-induced p65 and NF-κB activity in microglial cells.

<p>(A) Representative images of NF-κB p65 in microglial cells of each group. Cells were pretreated with or without PQQ for 1 h followed by 100 ng/ml LPS treatment for 2 h. Microglial cells were incubated with NF-κB p65 antibody and immunofluorescence microscopy was used to visualize the localization of NF-κB p65 (Green; a-c), boxed regions in (a-c) are also shown at×200 (j-m). Nuclei were visualized using DAPI counterstaining (Blue; d-f). (B) Cells were treated with 100 ng/m LPS for indicated time. p65 protein level was measured by western blot analysis. Non-phosphorylated p65 was used as loading control, and the expression of p-p65 was normalized to control and quantified by densitometric analysis. The results shown are mean ± S.E.M. of three independent experiments. <i>*p<0.05 vs. control group, #p<0.05 vs. only LPS group.</i></p