生物制药领域蛋白质团聚检测技术的研究进展

21世纪是生物制药的黄金时代,以重组蛋白药物和治疗性抗体为代表的蛋白质药物已成为生物技术药物的重要组成部分,其对恶性肿瘤、自身免疫性疾病、病毒感染等重大疾病的治疗效果和生物安全性远高于小分子化学药物。但是蛋白质稳定性差,在生产、纯化、运输和储存等过程中,容易因环境刺激或自身稳定性等因素产生团聚,从而导致药效降低,并可能引发免疫反应。发展灵敏度高、分辨率好、简便实用的蛋白质团聚检测技术和方法,对蛋白质药物的研发和改良可起到积极的推动作用。本文针对蛋白质团聚的检测技术和方法进行综述,从检测原理、性能和应用范围等方面,对体积排阻色谱法、凝胶电泳法、分析超速离心法、场流分离法、浊度法、动态光散射法、纳米颗粒跟踪分析技术、流式细胞术和电子显微技术等进行分析比较,并对蛋白质团聚检测技术未来的发展趋势进行了展望。国家自然科学基金项目(Nos.21225523,21627811)资助~

贺黄本立院士九十华诞

黄本立院士,1925年9月21日出生于香港,祖籍广东新会;光谱化学家,厦门大学教授。1945年—1949年就读于岭南大学。1950年—1986年任职于中国科学院长春应用化学研究所,历任助理研究员、副研究员、研究员;1986年调厦门大学任教至今。曾任中国化学会理事长、中国光谱学会副理事长、分析化学学科委员会主任委员、《光谱学与光谱分析》主编,《分析化学》、《分析科学学报》、《分析试验室》、《冶金分析》、《化学进展》等十多种国内期刊编委或顾问,SPECTrOCHIM

化学生物学专业实验室创建实践与体会

随着知识经济的高速发展,新兴、交叉、边缘学科不断涌现,交叉学科专业实验室建设成为高等院校教学改革中的重要课题。该文主要从实验室创建前科学调研、创新观念合理规划、实验教学大纲与教材编写和实验室的软硬件建设等方面,简要阐述了化学生物学专业实验室的创建实践与体会。国家基础科学人才培养基金项目(J1310024);;2014年福建省高等学校教学改革研究专项(JAS14634

The Establishment of Using Liquid Chip Technology to Quantify Prostate Specific Antigen in Human Serum

目的建立利用液相芯片技术定量检测人血清中前列腺特异性抗原的反应模式,并对该方法进行评价。方法应用液相芯片技术检测50例前列腺增生患者血清中的前列腺特异性抗原(PSA),评价该方法的线性范围、最低检测限、精密度等指标,并将结果与化学发光免疫分析法(ClIA)进行相关性分析。结果液相芯片技术检测PSA标准曲线的线性范围为0.022~129.6ng/Ml,PSA的最低检测限为0.018ng/Ml,检测PSA的批内CV为2.18%~2.28%,批间CV为1.61%~4.18%。用液相芯片技术和化学发光法分别对受检血清标本进行PSA定量分析,结果表明两法差异无显著性(P>0.05),r=0.9984,相关性良好。结论液相芯片技术具有线性范围广、灵敏度高、重复性好、节省样品和时间等优点,具有临床应用潜力。Objective To quantify prostate specific antigen in human serum using liquid chip method and evaluate the performance of the method.Methods Liquid chip method was employed to detect the PSA in 50 cases of benign prostatic hyperplasia patients'serum,evaluating the index of linear range,detection limit and accuracy,and make a correlation analysis with chemiluminescence immunoassay(CLIA).Results The linear range for the measurement of PSA was 0.022--129.6ng/mL and the lowest detection limit for PSA was 0.018 ng/ml.The intra-and inter-assay coefficients of variation were 2.18%～2.28% and 1.61%～4.18% respectively.Liquid chip method and CLIA were used to quantify PSA in human serum.The results showed that there were no significant differences in these two methods(P>0.05),but they had a good correlation(r=0.9984).Conclusion Liquid chip immunoassay has wide linear range,high sensitivity,repeatability,saving time and samples,with potential clinical applications

Preparation of fluorescence-encoded microspheres in a core-shell structure for suspension arrays

Fluorescence-encoded microspheres are widely used in the detection and analysis of biological molecules, especially in suspension arrays. Here, we report an efficient strategy for the preparation of fluorescence-encoded polystyrene microspheres with desirable optical and surface properties. The micron-sized, monodisperse polystyrene seed beads were first synthesized by dispersion polymerization. Then, dye molecules and carboxyl functional groups were copolymerized on the surface of the seed beads by forming a core-shell structure. Rhodamine 6G (R6G) was used as a model dye molecule to prepare the fluorescent beads, and the fluorescence intensity of the beads can be precisely controlled by adjusting the quantity of R6G. These fluorescent beads were characterized by environmental scanning electron microscopy, laser scanning confocal microscopy, and spectrofluorometry. The differences of the fluorescence spectra between fluorescent beads and R6G in solution were investigated. Twelve kinds of fluorescent beads encoded with different R6G fluorescence intensities were prepared, and they can be clearly distinguished on a conventional flow cytometer. Furthermore, the encoded beads are stable in water and resistant to photobleaching, which is crucial for their potential applications in diagnostic assays and imaging. Detection of human alpha fetoprotein antigen via a sandwich microsphere-based immunoassay yielded a detection limit of 80 pg mL(-1), demonstrating that the fluorescence-encoded microspheres synthesized herein are efficient in serving as the microcarriers in suspension arrays. As both the encoding and functionalizing procedures are made simultaneously, the newly designed technique is extremely simple and time-saving. Moreover, it could be readily applicable to the preparation of a wide size range of fluorescent particles made by polymerization.National Natural Science Foundation of China [20675070]; Program for New Century Excellent Talents in University [NCET-07-0729]; NFFTBS [J0630429]; Scientific Research Foundation ; State Education Ministr

Beryllium uptake and related biological effects studied in THP-1 differentiated macrophages

Investigation of cellular uptake of metal compounds is important in understanding metal-related toxicity and diseases. Inhalation of beryllium aerosols can cause chronic beryllium disease, a progressive, granulomatous fibrosis of the lung. Studies in laboratory animals and cultured animal cells indicate that alveolar macrophages take up beryllium compounds and participate in a hypersensitivity immune response to a beryllium-containing antigen. In the present work, human monocyte cell line THP-1 was induced with phorbol myristate acetate to differentiate into a macrophage. This cell with characteristics of human alveolar macrophages was employed to study cellular beryllium uptake and related biological effects. Morphological changes, phagocytosis of fluorescent latex beads, and cell surface CD14 expression were used to verify the successful differentiation of THP-1 monocytes into macrophages. An improved mass spectrometry method for quantitative analysis of intracellular beryllium as opposed to the traditional radioisotopic approach was developed using ICP-MS. The influence of the solubility of beryllium compounds, exposure duration, and beryllium concentration on the incorporation of beryllium was studied. Our data indicated that the uptake of particulate BeO was much more significant than that of soluble BeSO4, suggesting the major cellular uptake pathway is phagocytosis. Nevertheless, subsequent DAPI nuclear staining and PARP cleavage study indicated that beryllium uptake had a negligible effect on the apoptosis of THP-1 macrophages compared to the unstimulated macrophage control. Meanwhile, no substantial variation of tumour necrosis factor-alpha production was observed for THP-1 macrophages upon beryllium exposure. These data imply alveolar macrophages could have some level of tolerance to beryllium and this may explain why most Be-exposed individuals remain healthy throughout life.Program for New Century Excellent Talents in University [NCET-07-0729]; Scientific Research Foundation for Returned Overseas Chinese Scholars, State Education Ministr

Progress in the development of techniques based on light scattering for single nanoparticle detection

Nanoparticles have recently attracted extensive attention in view of their great potential in biomedicine and bioanalytical applications. Single particle detection via light scattering offers a simple and efficient approach for the size, size distribution, and concentration analysis of nanoparticles. In particular, intrinsic heterogeneity or rare events masked by ensemble averaging can be revealed. However, the sixth power dependence of Rayleigh scattering on particle size makes it very challenging to detect individual nanoparticles of small sizes. This article is intended to provide an overview of recent progress in the development of techniques based on light scattering for the detection of single nanoparticles.National Natural Science Foundation of China[20675070, 20975087, 90913015, 21027010]; Program for New Century Excellent Talents in University[NCET-07-0729]; Research Fund for the Doctoral Program of Higher Education of China[20090121120008, 20090121110009]; National Fund for Fostering Talents of Basic Science[J1030415

Langmuir probe potential measurements for reduced-pressure inductively coupled plasma mass spectrometry

A floating Langmuir probe was used to measure the apparent de offset potential of a reduced-pressure inductively coupled plasma near a substitute sampling orifice of a mass spectrometer. The experimental results demonstrate that the de offset potential causes the secondary discharge at the sampling orifice. The plasma potential is in the range +3.5 to +20 V and varies with the plasma operating conditions. The manner by which a water-cooled torch is shielded has a substantial effect on the plasma potential, then the secondary discharge. The measured values of the potential give a good explanation for the enhanced capacitive coupling effect in reduced-pressure ICP-MS reported previously

单个纳米颗粒的光散射检测技术进展

纳米颗粒因其在生物医学和生物分析领域具有重要的应用前景而备受关注.单个纳米颗粒的光散射检测技术是一种简单、有效地对纳米颗粒的尺寸、尺寸分布及浓度等进行表征的分析方法,尤其在揭露纳米颗粒的内在异质性方面具有独特优势.然而瑞利散射强度随粒径减小呈六次方衰减,使得小尺寸单个纳米颗粒的检测非常具有挑战性.本文对近年发展起来的多种单个纳米颗粒的光散射检测技术进行综述

多道ICP-AES与瞬时进样技术在线联用的信号采集和处理

报道了一种适用于多道ICP-AES和多种瞬时进样技术在线联用的信号采集、处理方法及相应软件.该方法适用范围较宽,已成功地用于4种不同联用技术的信号检测