拉曼光谱技术在恶性骨肿瘤患者红细胞中的研究应用

对单晶态血红蛋白和红细胞的拉曼光谱进行了研究.发现在600～1 750 cm-1的范围内,红细胞与单晶态血红蛋白的拉曼谱图极其相似,只有小量的峰移和加宽.实验结果表明可以通过分析红细胞的拉曼谱图中与heme相关的特征峰来研究红细胞内血红蛋白的结构及功能.对正常人和恶性骨肿瘤患者红细胞的拉曼光谱进行了分析,发现恶性骨肿瘤...     

M?ssbauer studies of hemoglobin in erythrocytes exposed to neutron radiation

We studied radiation effects on the stability of various states of hemoglobin (Hb) in red blood cells (RBC) irradiated with a very low dose of neutron rays, 50?μGy. We investigated RBCs isolated from blood of healthy donors. M?ssbauer spectroscopy was applied to monitor different forms of Hb. Our results show, for the first time, that oxyhemoglobin (OxyHb) and deoxyhemoglobin (DeoxyHb) are two Hb forms sensitive to such a low neutron radiation. Both Hbs change into a new Hb form (Hb_irr). Additionally, OxyHb transfers into HbOH/H₂O, which under our experimental conditions is resistant to the action of neutron rays.     

Analysis of blood proteins on protein arrays with a spectral surface plasmon resonance biosensor

We investigated whether blood proteins can be analyzed on protein arrays with a spectral surface plasmon resonance (SPR) biosensor. We modified gold arrays with a mixture of mercaptoundecanoic acid and mercaptohexanol, and immobilized blood proteins-hemoglobin (Hb) and haptoglobin (Hp), or their antibodies. We analyzed interactions of Hb with Hp on protein arrays by measuring the shift of resonance wavelength, and a significant interaction was observed when Hp was used as a ligand. Then, we fabricated antibody arrays with antibodies against Hb and Hp, and analyzed binding of blood proteins onto the arrays. Anti-Hb showed a specific interaction with Hb on the antibody arrays. Interaction of anti-Hp with Hp was specific when Hp was used as capture molecules. The shift of resonance wavelength caused by the interaction of blood proteins was explained by changes of refractive index on the gold surfaces of protein arrays. The present work suggests that spectral SPR biosensors can be used as a useful tool for the analysis of blood proteins on protein arrays.     

激光诱导血红蛋白分子的多光子荧光

本文首次报道了血红蛋白吸收二个和四个1.06μm的光子后所发射的荧光特性,并分析了其产生的原因。     

The effect of some medical treatments of thalassemia on the red blood cells

The Mössbauer spectroscopy (MS) and circular dichroism (CD) measurements have been used to investigate the effect of some medical treatments on the red blood cells (RBCs) of the patients with HbH discase and β-thalassemia (Thal.) major, respectively. The results indicate that both splenectomy and treatment with myleran are effective to alleviate the symptoms of anemia for some patients, but both of them are different in the effect on the RBCs of the patients. On the basis of the results, a hypothesis on the course of denaturation in hemoglobin (Hb) of the patients is proposed.     

Ⅱ型糖尿病人与大鼠红细胞拉曼光谱学比较

使用激光共聚焦拉曼光谱仪测量正常大鼠红细胞、正常人红细胞、糖尿病STZ造模大鼠红细胞、糖尿病四氧嘧啶造模大鼠红细胞和人Ⅱ型糖尿病红细胞的拉曼光谱,应用主成分分析(principal component analysis,PCA)结合支持向量机(support vector machines,SVM)分类器对数据进行判别分析,然后采用类间距离判断两种造模方法与人Ⅱ型糖尿病的接近程度。结果发现糖尿病红细胞与正常红细胞的拉曼光谱存在明显差异,糖尿病在酰胺 ⅥCO变形振动谱带处峰高显著,并在酰胺ⅤN—H变形振动谱带处谱线出现偏移,属于磷脂的脂酰基C—C骨架1 130 cm-1谱线增强,1 088 cm-1谱线强度减弱,说明糖尿病红细胞膜的通透性增强。PCA结合SVM可以很好地区分以上5类红细胞的拉曼光谱,分类器测试结果表明分类准确度达100%。通过分别计算两种造模方法与人Ⅱ型糖尿病的类间距离,发现STZ造模法更接近人Ⅱ型糖尿病。由此得出结论：拉曼光谱法可以用于糖尿病诊断,大鼠糖尿病STZ造模法更接近人类Ⅱ型糖尿病。     

基于同步移相显微干涉的血红细胞形貌测量方法

血红细胞的形貌特征是医学领域对多种疾病进行预防和诊断的一项重要指标,提出一种同步移相显微干涉法实现对血红细胞形貌的动态测量。搭建了透射式显微干涉成像系统,测量了100 μm内径模拟微血管内、名义直径为7 μm～8 μm、高度最大值为2 μm的新西兰兔血红细胞,针对血红细胞所处的微血管环境提出了基于微血管相位相减的血红细胞形貌提取方法和成像放大率校正方法,实验得到模拟微血管内的血红细胞平均直径7.757 μm和平均最高高度2.022 μm,验证了本方法具有在体定量测量血红细胞形貌的潜力。     

MRI of thermally denatured blood: methemoglobin formation and relaxation effects

Focal regions of T₁-shortening have been observed in magnetic resonance imaging (MRI)-monitored thermal ablations of perfused tissues. The aims of this study were two-fold: to find evidence for heat-induced conversion of hemoglobin (Hb) to methemoglobin (mHb), and to investigate the effects of heat treatment of in-vitro blood components upon their MR relaxation times. Spectrophotometric studies were performed to confirm the heat-induced formation of methemoglobin. Preparations of whole and fractionated blood, previously submitted to elevated temperatures of 40°C to 80°C, were imaged and the relaxation times were calculated. Optical absorption spectra of samples containing free Hb, heated to 60°C, showed increased light absorption at 630 nm, evident of mHb presence. Short T₁ values in whole blood (1.13 s) and packed red blood cell (0.65 s) compartments, heated at 60°C, compared to their baseline values (1.62 s and 0.83 s, respectively), were attributed to mHb formation. In relation to MRI-guided thermal interventions, these results suggest a possible explanation for observation of hyperintense regions on T₁-weighted images.     

Intracellular sugars improve survival of human red blood cells cryopreserved at -80 degrees C in the presence of polyvinyl pyrrolidone and human serum albumin

Cryopreservation with impermeable protectants has great significance on storage of human red blood cells. It has become feasible to use glycerol free cryopreservation for human red blood cells. This study focuses on the effect of intracellular trehalose or glucose on human red blood cells cryopreserved in the presence of polymer. Red blood cells were cryopreserved for 48 h-72 h at -80 degrees C. The data showed that the loading efficiency of glucose was significantly higher than that of trehalose, but trehalose loading process induced more hemolysis than glucose loading process. Compared with the other groups, the combination of intracellular glucose, PVP, and human serum albumin can significantly decrease the percent hemolysis after cryopreservation (P<0.01). However, the percent hemolysis induced by intracellular trehalose was less than that induced by extracellular trehalose, but the difference was not significant (P<0.05). The adenosine 5'-triphosphate (ATP) level and 2,3-diphosphoglycerate (2,3-DPG) level of cryopreserved red blood cells were significantly less than those of fresh red blood cells. However, sugars can provide certain protection for ATP and 2, 3-DPG compared with red blood cells cryopreserved in the absence of sugars. The protection of glucose on the metabolic function was more than that of trehalose. Cryopreservation can increase the percentage of cells with exposed phosphatidylserine (PS), but the ability of trehalose to maintain PS normal distribution is higher than that of glucose. Furthermore, intracellular sugars can protect membrane integrity of cryopreserved red blood cells, although a small portion of cells appeared spherocytic or echinocytic shape. Finally, most membrane proteins of cryopreserved red blood cells were similar to the membrane proteins of fresh red blood cells, but trehalose can result in loss of glyceraldehyde phosphate dehydrogenase (GAPD) and peroxiredoxin 2. In conclusion, it is feasible to cryopreserve red blood cells using polymer, human albumin and sugars as main protectants. The cryoprotective effect of glucose may be better than that of trehalose in the presence of PVP and human serum albumin, because sugar loading process causes more cell injuries in case of trehalose compared to glucose, and these injuries in turn manifest themselves during subsequent cryopreservation and thawing. In the future, finding an approach to decrease the injuries during trehalose loading process still is critical.     

Researching on the solution variation of double-concave disc-shaped vesicle of alcoholism blood cells

The physical model of the interaction between ethanol and blood cells has been established by the biomembrane surface elasticity theory. Under the universal equation of double-concave disc-shaped bubble, the analytical solution has been deduced. It confirmed theoretically that ethanol could lead to the larger volume of red blood cells. The fluorescence experiment result verified this conclusion. When ethanol goes into the red blood cells, the changing osmotic pressure causes cells’ vertices moving to the center, and double-concave disc surface will be the spherical one. It results in the larger volume of the red blood cell and smaller surface area, and thus causes the ratio of the surface area to volume decrease significantly. So the resistance tensile capacity of red blood cells reduced. Alcoholism can cause blood viscosity and the clinical test result coincide this conclusion. This study can give certain reference for the research of mutual function mechanism between ethanol and blood cells.     

Healthy and unhealthy red blood cell detection in human blood smears using neural networks

One of the most common diseases that affect human red blood cells (RBCs) is anaemia. To diagnose anaemia, the following methods are typically employed: an identification process that is based on measuring the level of haemoglobin and the classification of RBCs based on a microscopic examination in blood smears. This paper presents a proposed algorithm for detecting and counting three types of anaemia-infected red blood cells in a microscopic coloured image using circular Hough transform and morphological tools. Anaemia cells include sickle, elliptocytosis, microsite cells and cells with unknown shapes. Additionally, the resulting data from the detection process have been analysed by a prevalent data analysis technique: the neural network. The experimental results for this model have demonstrated high accuracy for analysing healthy/unhealthy cells. This algorithm has achieved a maximum detection of approximately 97.8% of all cells in 21 microscopic images. Effectiveness rates of 100%, 98%, 100%, and 99.3% have been achieved using neural networks for sickle cells, elliptocytosis cells, microsite cells and cells with unknown shapes, respectively.     

基于拉曼光谱方法研究不同剪切力下的红细胞亚损伤

利用共聚焦拉曼光谱技术,对人工心脏泵不同剪切应力下受到亚损伤的红细胞进行实验研究,验证拉曼光谱对红细胞亚损伤程度的评估能力,为血液损伤评价提供了一种新的思路。实验采集血红蛋白和红细胞的拉曼光谱标准谱图并进行对比分析,以确定红细胞谱图特征峰的归属。用血液剪切力试验平台对测试血样施加暴露时间为1 s,大小分别为0,50,100,150,200,250和300 pa的剪切力。利用共聚焦拉曼仪器,在10倍长焦物镜,532 nm激光光源波长,积分时间10 s,积分次数2次,2.5 mW功率下采集剪切应力作用后的红细胞拉曼谱图。通过归一化的方法对比红细胞的拉曼谱图变化,评估红细胞亚损伤的程度,运用曲线拟合方法对特征峰和剪切应力进行拟合,验证拉曼光谱对红细胞亚损伤的评估能力。对比血红蛋白和红细胞的拉曼光谱标准谱图发现,红细胞谱图能够反映血红蛋白的内部结构。且结果表明,拉曼光谱法可以用于区分不同剪切应力下亚损伤的红细胞,推断剪切应力可以透过细胞膜从而影响到其内部的血红蛋白结构。且随着剪切力的增大,1 376 cm-1位置左侧谱线呈现明显抬高趋势,1 549和1 604 cm-1位置的峰强增高,1 639 cm-1位置的氧浓度标记带ν₁₀振动谱带减弱。其中1 549 cm-1位置的峰强为亚铁离子高自旋带,在不同剪切力的作用下,峰强差异表现最明显,与剪切应力呈明显的正向线性关系,拟合效果良好。拉曼光谱法检测样本处理简单、耗时短、操作简便、重现性好,且可以精确的检测到细胞内部结构的细微变化,可以评估红细胞的亚损伤程度,弥补了传统评价溶血的方法的不足,为人工心脏泵血液损伤评价提供了新的技术手段,拓宽了拉曼检测方法的应用领域。     

流动注射化学发光法灵敏测定人血液和血清中血红蛋白

在碱性介质中,发现血红蛋白对luminol-hydrazine弱化学发光体系有显著的增强作用。结合流动注射法提出了luminol-hydrazine化学发光(FI-CL)测定血红蛋白的新方法,优化了化学发光检测条件和FI-CL系统的运行参数,考察了实际样品中可能含有的共存物质对血红蛋白测定的影响。结果表明,血红蛋白的线性范围为5.0×10-9～6.0×10-5 g·mL-1,检出限是5.8×10-10 g·mL-1 (9.0×10-12 mol·L-1),分别对5.0×10-7和3.0×10-6 g·mL-1的血红蛋白标准溶液平行八次测定所得RSDs分别为1.6%和1.5%。将提出的方法成功用于人血液和血清中血红蛋白含量的测定,加标回收率在83.0%～101.0%范围。结合化学发光光谱和紫外可见吸收光谱对血红蛋白催化luminol-hydrazine化学发光反应的机理进行了探讨。提出的方法具有灵敏度高、无需标记、操作简便、快速、易实现自动化操作等优点。     

NMR Water Proton T1 Mechanism in Blood Diluted by its Own Plasma

The water proton T1 in human blood diluted by its own plasma was measured with a FT-NMR spectrometer operating at 60MHz for protons. A linear relationship (with a correlation of 0.99) was found between the 1/T1 and hemoglobin content(Hb) in the blood. The exchange of water between the extracellular plasma and the intracellular Hb in blood is known to satisfy the fast chemical exchange conditions, and the decay of magnetization in blood is reported to have a single exponential. Therefore, the obtained relationship should represent fast chemfcal intracellular Hb and the extrace exchange between the lular plasma.     

Effective removal of adhering cells via ultrashort laser pulses

A technique for accurately and effectively removing adhering red blood cells in a blood plasma thin film via a picosecond pulsed laser was developed. The laser beam was focused to the surface of the film to generate plasma-mediated ablation and an automated stage was employed for raster scan. The SEM images showed that the red blood cells distributed in the ablation scanned area were removed neatly, leaving the surroundings and the film base intact. For cells across the boundary between the ablated and untreated areas, a trim cutting interface was observed. Complete ablation of red blood cells in the target area is achieved without visible thermal and collateral damage in the remaining structure. The removal method is very effective because it is not necessary to selectively focus a laser beam on individual target cells and remove cells one by one. The ablation is scanned over a certain size of area, enabling practical cell killing or microbial decontamination in clinical/industrial scale.     

血红蛋白与铜(Ⅱ)-茜素红S络合物相互作用的研究

采用UV-Vis光谱法,研究在pH 4.2的H₃PO₄-KH₂PO₄缓冲溶液中血红蛋白(Hb)与铜(Ⅱ)-茜素红S(ARS)络合物的反应。结果表明：Hb与Cu(Ⅱ)-ARS络合物相互作用形成红色的离子缔合复合物,该复合物的最大吸收波长为537 nm。在此波长下测得复合物的组成为n_Hb∶n_Cu(Ⅱ)∶n_ARS=1∶4∶8,表观摩尔吸光系数为1.52×105 L·mol-1·cm-1,Hb浓度在1.0×10-7～2.0×10-6 mol·L-1范围内与吸光度呈线性关系,回归方程为A=0.026 9+151 675ｃ(mol·L-1),相关系数r=0.997 2。考察了溶液酸度、试剂用量、反应时间与温度、离子强度、表面活性剂等因素对Hb-Cu(Ⅱ)-ARS复合物形成的影响,并对反应机理做了初步探讨,发现Hb与Cu(Ⅱ)-ARS络合物之间主要以静电引力相结合。进一步考察了常见氨基酸及金属离子对Hb-Cu(Ⅱ)-ARS复合物形成的影响。     

Deformation of Two-Dimensional Nonuniform-Membrane Red Blood Cells Simulated by a Lattice

To study two-dimensional red blood cells deforming in a shear flow with the membrane nonuniform on the rigidity and mass, the membrane is discretized into equilength segments. The fluid inside and outside the red blood cell is simulated by the D2Q9 lattice Boltzmann model and the hydrodynamic forces exerted on the membrane from the inner and outer of the red blood cell are calculated by a stress-integration method. Through the global deviation from the curvature of uniform-membrane, we find that when the membrane is nonuniform on the rigidity, the deviation first decreases with the time increases and implies that the terminal profile of the red blood cell is static. To a red blood cell with the mass nonuniform on the membrane, the deviation becomes more large, and the mass distribution affects the profile of the two sides of the flattened red blood cell in a shear flow.     

Polarization Raman study of protein ordering by controllable RBC deformation

Polarized Raman spectroscopy is used to provide evidence of hemoglobin protein ordering as the red blood cell (RBC) is stretched with optical tweezers. The stretching of the cell is intended to mimic the deformation that it experiences as it travels through vessels and capillaries. The depolarization ratios for a number of heme Raman bands change as the cell is stretched, confirming the semi‐ordered nature of the hemoglobin ensemble in the cytoplasm. Furthermore, trends observed in the ratio shifts point to increased packing and ordering of the Hb after cell stretching. This evidence should shed more light on to the role of deformation in the RBC function. Copyright © 2009 John Wiley & Sons, Ltd.     

血液中碳氧血红蛋白饱和度的双波长分光光度简便测定法

本文报道血液中碳氧血红蛋白基于吸光度差值的双波长分光光度检测方法,检血用含有Na2S2O4的溶液释得到HbCO和Hb的双组分溶液,Hb有相同吸光度的双波长处的吸光度差值只反映HbCO的浓度,HbCO有相同吸光度的双波长处的光度差值只反映Hb的浓度,测定了这此吸光度差值可以利用约定的公式计算出检血的HbCO%.进行比较实验,当检出是由HbCO和Hb溶液混合制备时,本方法测定值与理论值严格一致,当检血是中毒案件检血时,本方法测定值与可靠方法的测定值严格一致.     

Directed energy transfer due to orientational broadening of energy levels in photosynthetic pigment solutions

The directed non-radiative energy transfer through monomeric molecules of chlorophyll “a” and pheophytin “a” at high concentrations (c~10^-2 M) in a rigid matrix of polyvinylbutyral has been found by using the nanosecond laser spectrofluorimeter. The phenomenon is caused by orientational broadening of pigment molecular spectra owing to its interaction with a solvent. The observed temporal shift of the luminescence spectrum to the red region in a nanosecond time scale as well as the red shift of the time integrated spectrum at a high concentration of pigment molecules and the monotonic growth of the luminescence lifetime with a shift to the red region of the spectrum served as indications of the directed energy transfer in the sample. The non-radiative energy transfer from monomeric molecules towards aggregates is also directly demonstrated by the deformation of instantaneous luminescence spectra in the long-wavelength range (λ>700 nm). The role and the possibility of the directed energy transfer between molecules with orientationally broadened spectra in the biological systens are discussed.