Preparation and identification of multifunctional mesoporous silica nanoparticles for in vitro and in vivo dual-mode imaging,theranostics, and targeted tracking

Mesoporous silica nanoparticles (MSNs) can provide a structural foundation for a new generation of nanocarriers with a broad range of functionalities. Multifunctional MSNs can serve as all-in-one diagnostic and therapeutic tools that can be used to simultaneously visualize and treat various diseases, such as cancer. This research study is the first time that two lanthanide-based imaging systems have been combined to incorporate controlled drug release and targeted tracing into a single MSN-based nano-platform for a novel theranostic drug delivery system. Doping lanthanide ions, i.e., europium (Eu) and gadolinium (Gd) ions, into an MSN structure (EuGd-MSNs) imparts fluorescence and magnetism to the nanostructure that can be used to develop magnetic resonance imaging (MRI) and biological fluorescence tools. Current cancer research has revealed that most human cancer cells express a large number of folate receptors on their surface. Grafting folic acid (FA) onto the EuGd-MSN surface (EuGd-FA-MSNs) imparts a targeting function to the MSN because of the specificity of the binding of FA to cell surface receptors. Furthermore, grafting anticancer drugs, such as camptothecin (CPT), onto the surface of these MSNs by forming disulfide bonds (EuGd-SS-CPT-FA-MSNs) enables intracellular controlled drug release. A high concentration of intracellular glutathione cleaves the disulfide bond to release the drug and treat the disease. The results of in vitro and in vivo studies show that the functionalized MSNs can be successfully used as a platform to integrate dual-imaging, targeting, and therapeutic treatment in multifunctional diagnosis drug delivery systems.     

荧光原位杂交法快速鉴定血培养阳性葡萄球菌的临床应用

目的快速鉴定血培养中的金黄色葡萄球菌和凝固酶阴性葡萄球菌(CoNS),结合临床快速判定是否为污染菌。方法采用荧光原位杂交法鉴定血培养中的金黄色葡萄球菌和CoNS,杂交结果若为CoNS,根据临床资料进行判断,并与文献推荐的污染判断法进行结果比较。结果探针的特异性经由标准菌株和临床分离菌株证实。金黄色葡萄球菌探针的特异性和敏感性均为100%,GoNS探针的特异性和敏感性分别为100%和95.5%。179株CoNS中117株判断为污染菌,污染率为68%,与文献推荐的污染判断方法一致。结论荧光原位杂交法适用于血培养中的金黄色葡萄球菌和CoNS的快速鉴定,以排除CoNS污染。     

On the physical properties of red cell ghost membranes in the affective disorders and psychoses A fluorescence polarization study

Membrane order was measured in the erythrocyte ghost membranes of DSM-III schizophreniform disorder (SF), DSM-III schizophrenic (SCZ) and DSM-III manic (bipolar) (M) patients and a group of age- and sex-matched controls. Fluorescence polarization with the probe 1,6-diphenyl1-1,3,5-hexatriene was used to determine the steady-state fluorescence anisotropy (r_s). The SF group showed a significant increase in r_s (Δr_s = 0.037) from the control group. Although the means were not significantly different, 3 of 8 Ms and 5 of 8 SCZs also had r_s values > the highest control value. Thermotropic behavior of the membranes was evaluated over the range of 40 to 20°C. No difference among groups in membrane enthalpy was detected. Thus, the differences in r_s appear to be associated with differences in entropy. Phosphatidylcholine (PC) levels, which were known to be abnormal in these patients, were compared with the r_s values. A significant (P < 0.001, R= -0.63) linear correlation between r_s and membrane PC levels was observed. Overall these data further support the view that unusual membrane biophysical factors may occur with high frequency in the psychoses and affective disorders.     

早期糖尿病视网膜病变的视网膜电图分析

目的 分析早期糖尿病视网膜病变(DR)的闪光视网膜电图(F—ERG)和视网膜电图震荡电位(OPs)各参数的变化特点，寻找反映早期DR的敏感指标。方法 对16例(32只眼)正常人进行OPs和F-ERG检测。对27例(53只眼)糖尿病病人进行眼底荧光血管造影(FFA)、OPs和F—ERG检测。结果 OPs中Os波幅、O4波幅、OPs总波幅及F-ERG中b波峰潜时较其他指标敏感，其中O4波幅和b波峰潜时为最敏感指标，但均不能反映早期DR的严重程度。结论 OPs的O4波幅和F—ERG的b波峰潜时可作为早期糖尿病视网膜病变诊断的敏感指标。     

In vivo quantification of fluorescent molecular markers in real-time: A review to evaluate the performance of five existing methods

With the advent of molecular-targeted fluorescent markers, there is a renewed interest in fluorescence quantification methods that are based on continuous wave excitation and multi-spectral image acquisition. However, little is known about their in vivo quantification performance. We reviewed the performance of five selected methods by analytically describing these and varying input parameters of irradiance, excitation geometry, collection efficiency, autofluorescence, melanin content, blood volume, blood oxygenation and tissue scattering using optical properties representing those for human skin. We identified one method that corrects for variations in all parameters. This requires image acquisition before and after marker administration, under identical geometry. Hence, it is suited for applications where the site of interest can be relocated (e.g. anaesthetized animals and dermatology). For applications where relocation is not possible, we identified a second method where the uncertainty in the fluorescence signal was ±20%. Hence, use of these methods can substantially aid in vivo fluorescence quantification compared to use of the raw fluorescence signal, as this changed by more than 3 orders of magnitude. Since these methods can be computed in real-time, they are of particular interest for applications where direct feedback is critical, as diagnostic screening or image-guided surgery.     

Fluorescence behaviour of human arterial tissue

Total fluorescence from arterial tissue is influenced by three factors: the absorption coefficient of tissue at a specific excitation wavelength, the laser excitation power and the fluorescence coefficient which is related to chemical species in tissue. These various influences were demonstrated by the following experimental results in vitro: (1) the effect of increasing power on fluorescence intensity, (2) the total fluorescence intensity in normal aorta and plaque and (3) the effect of a chromophore such as β-carotene on total fluorescence intensity. The fluorescence intensity of normal artery is an incremental function of laser excitation power, and the fluorescence emission from normal artery compared to fluorescence emission from plaque is significantly different at the same excitation power. The total fluorescence of normal artery was measured to be twice as great as that of atheromatous plaque (relative mean ratio of 2.58±0.46 compared to unity,p<0.0002 at 488 nm; relative mean ratio of 2.57±0.51 compared to unity,p<0.0009 at 514 nm). The total fluorescence emission decreases with the increase of β-carotene content in arterial tissue (R=0.97). These emission differences, when intensified by an exogenous chromophore of β-carotene, may provide an improved guidance signal for diagnosis of plaque from normal artery during laser angioplasty procedures.     

荧光分光光度法测定水产品中烷烃类、芳烃类污染物

目的：对使用荧光分光光度法测定水产品中烷烃类、芳烃类污染物的方法进行验证,并检测三类样品。方法：通过氢氧化钠-乙醇皂化处理,用正已烷萃取,用石油醚溶解,测量萃取物荧光强度值进行定量。结果：测得标准油在0.00～12.0μg/ml范围内线性关系良好,相关系数0.9999,检出限为0.2 mg/kg,平均回收率90.2%,RSD5.4%。结论：本法准确度、灵敏度高,检出限低,操作稳定性好。     

荧光偏振检测HBV DNA C区BCP双突变

目的：建立简便、灵敏的HBV DNA序列中BCP双突变点的检测方法．方法：采用终点终止法和偏振光检测技术进行点突变的检测．首先对HBV C区基因进行PCR扩增，然后用特异探针与扩增产物中待测核苷酸的下游序列杂交，使探针的3’端可以在DNA聚合酶的催化下，依据其互补链上的待测核苷酸连接上一个标有特定荧光素的ddNTP，然后检测该3’端带有荧光素的探针，根据检测到的荧光素种类和偏振光的强度可以判定待测点是何种核苷酸．结果：该方法可以检测出HBV基因序列中BCP双突变核苷酸类型以及检测1个拷贝的模板，并且可以从BCP野性株DNA序列中检出5％BCP双突变DNA序列．结论：该技术可以检测血清中HBV DNA C区BCP双突变．     

Selective localisation of neuro-specific T lymphocytes in the central nervous system

Using experimental autoimmune encephalomyelitis (EAE) in the rat as a model of central nervous system (CNS) inflammation, activated and quiescent T lymphocytes with different antigen specificities were labelled with the fluorescent dye Hoechst 33342 and tested by fluorescence microscopy for their ability to accumulate in different regions of the spinal cord and in other organs at varying times post inoculation. With this highly sensitive assay it was found that activated myelin basic protein (MBP)-specific T cell lines accumulated in the spinal cord (a 1000-fold increase in the lumbar/sacral region by day 4) and caused clinical signs of EAE. In contrast, interleukin-2 (IL-2)-maintained (quiescent) MBP-specific T cell lines failed to accumulate in the CNS and cause disease. Activated ovalbumin (OA)-specific and purified protein derivative of tuberculin (PPD)-specific T cell lines were also found at significantly higher levels in the spinal cord than non-activated cells although they failed to accumulate to a substantial degree when injected alone. When injected with activated MBP-specific T cells the activated OA- and PPD-specific cell lines accumulated in the spinal cord following initial accumulation of the MBP-specific cells, demonstrating that during the inflammatory process there is considerable non-specific recruitment of cells into the inflammatory site. CNS accumulation of activated MBP-specific T cell lines occurred 1-2 days later in irradiated animals than in non-irradiated recipients. This was consistent with irradiated animals also exhibiting a later onset of disease and suggests that irradiation may directly affect the endothelium in a way that makes it less adhesive. In conclusion, this study demonstrates that activated lymphocytes of any specificity enter the spinal cord, and that the neuro-antigen specific cells accumulate there and lead to the recruitment of other cells. Non-activated cells, even those with neural antigen specificity fail to enter the cord. Understanding the nature of what an 'activated' lymphocyte is may allow us to design strategies to inhibit such immune-mediated inflammation.     

In vivo fluorescence spectroscopy and imaging of human skin tumours

The feasibility of using in vivo autofluorescence for the diagnosis of skin cancer was evaluated. In vivo fluorescence measurements were performed on healthy human volunteers, and patients with different types of benign and malignant skin tumours. Fluorescence spectra as well as fluorescence images were acquired. The excitation-emission matrix of normal skin (n=3) showed a broad peak at the shortest excitation wavelength (365 nm) and at 440 nm fluorescence wavelength, smoothly decreasing towards longer excitation and fluorescence wavelengths. Non-melanoma skin tumours (n=31) and control skin excited with 375 nm showed a broad fluorescence band from 400 to 700 nm, peaking around 436 nm. No significant differences in measurements between tumours and the corresponding control sites were found. A large spatial variation in the fluorescence intensity was observed both in the tumours and in the control sites. Standard deviations found ranged from 0.15 to 1.5 times the mean fluorescence. Fluorescence images, excited with 375 nm and taken with an image intensified CCD camera, on eight malignant melanomas and eight benign pigmented lesions did not indicate any fluorescence intensity distribution specific to the malignancy of the lesion. Neither the shape of the fluorescence spectra, nor the spatial distribution of the fluorescence intensity showed any signature specific to the histopathological nature of the lesions investigated. Optical diagnostics of skin tumours using the autofluorescence does not seem to be feasible at the present time.