一种基于危险信号的拒绝服务入侵检测方法

针对拒绝服务(Denial of Service,DoS)攻击的特点,提出了一种基于免疫危险理论的新型入侵检测方法,设计、实现了检测算法和抗体变异、进化算法。引入血亲类方法分类抗原/抗体,定义了抗原凋亡和坏死的过程,定量计算抗原危险信号和网络危险度,并以此检测DoS攻击。仿真实验表明该方法不仅具有基于传统人工免疫理论的入侵检测自学习、自适应的优点,而且误警率降低87.5%,检测效率更高。     

基于聚硫堇的一次性O型口蹄疫抗原酶免疫传感器的研制

将O型口蹄疫酶标抗体掺杂于有机-无机溶胶凝胶中,并修饰于聚硫菫的丝网印刷碳电极表面,从而制备一次性口蹄疫病毒抗原(FMDV-Ag)酶免疫传感器。试验中采用直接免疫分析法检测口蹄疫病毒抗原。根据免疫反应前后还原峰电流下降的百分率K值的大小实现对抗原的检测。在优化条件下,免疫传感器检测抗原的线性范围为77～388ng/mL,最低检出限为32ng/mL。免疫电极具有较好的特异性、重现性(RSD=5.6%)、稳定性(10d后电流响应为初始值的89.5%)和准确性(与AGP符合率为95%)。因此,该免疫传感器有望用于O型口蹄疫抗原的快速检测。     

Detection of urinary biomarkers for early diagnosis of acute renal allograft rejection by proteomic analysis

Acute allograft rejection has been recognized as a major impediment to improved success in renal transplantation. Timely detection and control of rejection are very important for the improvement in long-term renal allograft survival. Thus, biomarkers for early diagnosis of acute rejection are required urgently to clinical medication. This study seeks to search for such biomarker candidates by comparing patients' pre-treatment urinary protein profiling with their post-treatment urinary protein profiling. A total of 15 significantly and consistently down-regulated protein candidates were identified. Among them, alpha-1-antichymotrypsin precursor (AACT), tumor rejection antigen gp96 (GP96) and Zn-Alpha-2-Glycoprotein (ZAG) were selected for further analysis. The results indicated that Western Blot assay of AACT, GP96 and ZAG had advanced the diagnosis time of acute renal rejection by 3 days, compared with current standard clinical observation and laboratory examination. Furthermore, the double-blind detection revealed that the accuracy, sensitivity and specificity of the diagnosis of acute renal rejection of AACT, GP96 and ZAG were 66.67%/100%/60%, 83.33%/100%/80% and 66.67%/100%/60%, respectively, and 100%/100%/100% in combination. In conclusion, urinary protein AACT, GP96 and ZAG could be a set of potential biomarkers for early non-invasive diagnosis of the acute rejection after renal transplantation.     

Engineering of protein epitopes: a single deletion in a snake toxin generates full binding capacity to a previously unrecognized antibody

Structural features associated with the ability of a monoclonalantibody (mAb) to discriminate between protein variants areidentified and engineered. The variants are the curaremimetictoxin from Naja nigricollis and erabutoxin a or b from Laticaudasemifasciata which differ from each other by 16 substitutionsand one insertion. The neutralizing mAb M1 recognizes with highaffinity a topographical epitope on the surface of toxin , butfails to recognize the erabutoxins although they possess mostof the residues forming the presumed epitope. Examinations ofthe toxin and erabutoxin 3-D structures and molecular dynamicssimulations reveal several differences between the variants.In particular, the region involving the ß-turn 17–24is organized differently. Analysis of the differences foundin this region suggests that the insertion (or deletion) atposition 18 of the variant amino add sequences is particularlyimportant in determining the differential cross-reactivity.To test this proposal, residue 18 was deleted in one erabutoxinusing sitedirected mutagenesis, and the biological propertiesof the resulting mutant were examined. We found that full antigenicitywas restored in the previously unrecognized variant. The implicationsof this finding are discussed.     

Sensitive Detection of Carcinoembryonic Antigen Using Stability‐Limited Few‐Layer Black Phosphorus as an Electron Donor and a Reservoir

The instability of few‐layer black phosphorus (FL‐BP) hampers its further applications. Here, it can be demonstrated that the instability of FL‐BP can also be the advantage for application in biosensor. First, gold nanoparticle/FL‐BP (BP‐Au) hybrid is facilely synthesized by mixing Au precursor with FL‐BP. BP‐Au shows outstanding catalytic activity (K = 1120 s^?1 g^?1) and low activation energy (17.53 kJ mol^?1) for reducing 4‐nitrophenol, which is attributed to the electron‐reservoir and electron‐donor properties of FL‐BP, and synergistic interaction of Au nanoparticles and FL‐BP. Oxidation of FL‐BP after catalytic reaction is further confirmed by transmission electron microscope, X‐ray photoelectron spectroscopy, and zeta potentials. Second, the catalytic activity of BP‐Au can be reversibly switched from “inactive” to “active” upon treatment with antibody and antigen in solution, thus providing a versatile platform for label‐free colorimetric detection of biomarkers. The sensor shows a wide detection range (1 pg mL^?1 to –10 µg mL^?1), high sensitivity (0.20 pg mL^?1), and selectivity for detecting carcinoembryonic antigen (CEA). Finally, the biosensor has been used to detect CEA in colon and breast cancer clinical samples with satisfactory results. Therefore, the instability of BP can also be the advantage for application in detecting cancer biomarker in clinic.     

Multi-enzyme layer-by-layer assembly for dual amplified ultrasensitive electronic detection of cancer biomarkers

The preparation and use of multi-enzyme layer-by-layer (LBL) assembled single wall carbon nanotube (SWCNT) composite labels for amplified ultrasensitive electrochemical detection of a cancer biomarker is described. The target protein, carcinoembryonic antigen (CEA), is sandwiched between an electrode surface-confined capture anti-CEA antibody and the secondary signal anti-CEA/enzyme-LBL/SWCNT bioconjugate. The dual biocatalytic signal amplification for CEA monitoring is achieved through both the numerous enzymes loaded on the CNTs and redox-recycling of the enzymatic products in the presence of the secondary enzyme and the corresponding substrate. Our novel dramatic signal amplification strategy, with a detection limit of 0.04 pg mL⁻¹, shows about 2-4 orders of magnitude improvement in sensitivity for CEA detection compared with other universal signal amplified assays, which makes our signal amplification approach hold great potential applications in detection of ultra-trace protein biomarkers.     

Identification of nasopharyngeal carcinoma antigens that induce humoral immune response by proteomic analysis

A proteomics-based approach has been used to identify proteins that commonly elicit a humoral immune response in nasopharyngeal carcinoma (NPC). Sera from 19 newly diagnosed NPC patients and 19 healthy individuals were analyzed for IgG autoantibodies against NPC proteins resolved by 2-DE. Protein spots that exhibited selective reactivity with sera from NPC patients were identified by MS. Among nine identified proteins, cytokeratin 19 (CK19), Erb3 binding protein (EBP1), and Rho GDP dissociation inhibitor-beta (Rho-GDI-2) induced autoantibodies in more than 36.8% of NPC patients but not in healthy individuals. Furthermore, Western blot analysis and immunohistochemical staining were performed to determine the expression and localization of CK19, EBP1, and Rho-GDI-2 in NPC and normal nasopharyngeal mucosal tissues. Up-regulated CK19 and EBP1, but not Rho-GDI-2, were observed in NPC vs. normal tissue. Subcellular localization of the three proteins in NPC tissue was same as that in the normal tissue. Thus, overexpression of CK19 and EBP1 may be one of the mechanisms for their autoantibody development in NPC. To validate the findings of a proteomic analysis, occurrence of autoantibodies against these three proteins was detected by immunoprecipitation and Western blot analysis in additional 30 NPC patients, 23 other types of cancer patients and 20 healthy individuals. Results showed that frequency of autoantibodies against CK19, EBP1 and Rho-GDI-2 in NPC patients was significantly higher than that in other types of cancer patients and healthy individuals. We conclude that CK19, EBP1 and Rho-GDI-2 may have utility in NPC screening and diagnosis.     

人工免疫系统中人工抗原提呈细胞的设计

反向选择模型是人工免疫系统中的经典模型,但存在训练代价高、覆盖率低的问题.危险理论模型是人工免疫系统中新引进的模型,在降低训练代价,提高覆盖率上具有优势.建立人工免疫危险理论模型必须解决的关键问题之一是:在保持人工免疫系统自适应特征的前提下,设计人工抗原提呈细胞.本文从抗原提呈细胞的生物学原理出发,设计了人工免疫系统中的人工抗原提呈细胞,给出单个人工抗原提呈细胞的结构、细胞上Toll样受体的结构,以及人工抗原提呈细胞群体的结构;设计了人工抗原提呈细胞群体的生命周期.初步的实验结果证明所设计的人工抗原提呈细胞具有自适应发现恶意软件的能力.     

基于小生境技术的多样性抗体生成算法

新的入侵方法以及网络计算环境的不断变化,使得入侵越来越难以防范.因此应用人体免疫机制构建下一代入侵检测系统成为一个新的研究热点.本文采用一种新的抗原－抗体编码方法,并在此基础上,提出一种基于共享函数的小生境遗传算法,用来产生多样性的抗体基因,最后给出多样性评价函数以验证算法的有效性.实验结果表明:该算法能够使抗体在演化过程中保持较好的多样性.