Independency of Fe ions in hemoglobin on immunomagnetic reduction assay

Immunomagnetic reduction (IMR), which involves measuring the reduction in the ac magnetic susceptibility of magnetic reagents, is due to the association between bio-functionalized magnetic nanoparticles and target bio-molecules. This has been demonstrated for assaying proteins in solutions free of Fe ions, such as serum. In this work, the validity of IMR assay for samples rich in Fe ions like hemoglobin (Hb) is investigated. According to the results, there is no magnetic signal contributed by Fe-ion-rich Hb. Furthermore, the results show a high sensitivity in assaying hemoglobin A1c (HbA1c) by using IMR.     

Polarized neutron reflectometry from the interface of the heterostructures SiO2(Co)/Si and SiO2(Co)/GaAs

Polarized neutron reflectometry is used to investigate SiO₂(Co) granular films (70 at% of Co nanoparticles in SiO₂ matrix) deposited on Si and GaAs substrates. The aim of the study is to compare magnetization depth profiles in two systems: in SiO₂(Co)/GaAs heterostructure which shows at room temperature giant injection magnetoresistance (IMR) with the system SiO₂(Co)/Si which reveals almost no IMR effect. We found that at room temperature and at the same value of external magnetic field mean magnetization in the SiO₂(Co)/GaAs sample is much higher than in the case of SiO₂(Co)/Si. We also demonstrate that magnetic scattering length density, and hence, magnetization profile strongly depends on the substrate. We show that SiO₂(Co)/Si heterostructure is ferromagnetically ordered within the temperature range between 120 and 460 K what could explain a weak IMR.     

Analysis of ploidy and the patterns of amplified fragment length polymorphism and methylation sensitive amplified polymorphism in strawberry plants recovered from cryopreservation

Shoot-tips of 10 strawberry genotypes were successfully cryopreserved using a modified encapsulation-dehydration method. All genotypes survived cryopreservation with high survival and regeneration rates. Eight Joho single-bud sibling lines were established as a model system for genetic analysis. Although cytological examination found chromosomal variation in both non-cryopreserved and cryopreserved samples, the ploidy constitution remained relatively stable after cryopreservation. DNA samples digested with MseI and PstI were used for amplified fragmentation length polymorphism (AFLP) assay. In 16 primer combinations, only one, namely, PCCA-MCAG, detected one site where band pattern changed after cryopreservation, which might be contributed to the change in DNA methylation status at PstI recognition site. Methylation sensitive amplified polymorphism (MSAP) assay was carried out for further investigation on the influence of cryopreservation on DNA methylation status. It was found that cryopreservation induced a significant change in DNA methylation status.     

A fluorescence based assay for DNA damage induced by radiation,chemical mutagens and enzymes

A simple and rapid assay to detect DNA damage is reported. This novel assay is based on changes in melting/annealing behavior and facilitated using certain dyes that increase their fluorescence upon association with double stranded (ds)DNA. Damage caused by ultraviolet (UV) radiation, chemical mutagens or restriction enzymes produced an assay response. UV radiation at 254 nm (approximating UV-C) and 360 nm (approximating UV-A) were used to induce the damage in dsDNA. Chemical damage was induced using several compounds with known effects on nucleic acids. Restriction enzymes Hind III, Msp1, Sau 3A1 were used to cut the plasmid (pUC19) at specific sequences in addition to the non-specific endonuclease DNase I. The effects of these types of damage on repeated melting and annealing of dsDNA were observed in real time using several fluorescence indicator dyes. Low concentrations of dsDNA (between 10 and 100 ng/ml) and small volumes (20 μl) were required for this assay. Repeated measures yielded a coefficient of variation of 2% (CV%). In addition to measuring various DNA damaging agents, the potential application of this assay to study the efficiency of various sun blocking agents against UV-induced DNA damage is discussed.     

Millimeter-Wave Lange Coupler Design and Its Application in the Monolithic Image Rejection Mixer Design

This paper reports on the design of a Ka-band monolithic Lange coupler and its application in the monolithic fourth-harmonic image rejection mixer. Detailed design and analysis using Ansoft-HFSS simulator have been carried out. The simulated results of the Lange Coupler show the insert loss is better than −3.64 dB; the amplitude balance is less than 0.55 dB and the phase balance is less than 0.65° from the 90° phase difference over the 30 to 40 GHz frequency range. The Lange Coupler is employed in a monolithic image rejection mixer that is fabricated by a commercial 0.18-μm pseudomorphic high electron-mobility transistor (pHEMT) process. The chip size is 1.4 mm × 1.9 mm. The image rejection ratio (IMR) is from 15 to 34 dB in the RF frequency range of 30 to 40 GHz.     

Evaluation of damage in Pacific oyster (Crassostrea gigas) spermatozoa before and after cryopreservation using comet assay

We examined the applicability of the comet assay (single cell gel electrophoresis assay) to estimate the quality of frozen-thawed Pacific oyster (Crassostrea gigas) spermatozoa. Comet assay was performed on semen before and after cryopreservation followed by fluorescent staining with propidium iodide to assess DNA integrity. After cryopreservation, the percentage of spermatozoa with damaged DNA significantly increased, while only about half of the cells displayed intact DNA, even when protected with 10 percent DMSO. All the considered parameters (head length, head area, head intensity, total length, total area, total intensity, tail length percent, tail area percent, and tail intensity percent) were higher than the oyster sperm protected with 10 percent DMSO-artificial sea water after freezing and thawing. Only tail length percent, tail area percent, and tail intensity percent were increased significantly after cryopreservation. The tail length percent was found to be the most sensitive indicator of the cryopreservation-induced DNA damage. Our freeze-thawing procedure significantly affected oyster sperm DNA, as indicated by the reduced fertilization rate when frozen-thawed oyster sperm are used. Irreversible alteration of the genome may prevent fertilization or alter normal embryonic development. This study is the first to demonstrate that the comet assay is an inexpensive, rapid and sensitive method for determining DNA damage in Pacific oyster sperm quality assessments.     

痕量样品高灵敏度快速测量方法与便携式系统研究

为了实现高灵敏度、快速、准确鉴定痕量病原菌核酸样品,介绍了一种痕量核酸样品高灵敏度快速测量方法,并发展了一种微流控芯片核酸等温扩增实时分子诊断技术,研制了新型微纳体系生化反应载体芯片。通过表面惰性处理降低芯片表面对生物分子的吸附影响,构建一种大数值孔径、长工作距离的便携式共焦光学检测系统,有效消除背景荧光的影响,提高了检测灵敏度。在微纳升(7μL～40nL)试剂消耗反应体系水平,实现检测灵敏度5个DNA分子拷贝数,并以呼吸道感染疾病的大肠埃希氏菌检测为例,开展临床应用研究,满足低成本临床医疗应用需要。     

A generalized time-domain model for nonlinear inter-satellite microwave photonics links under dual-tone modulation

A generalized time-domain model is presented for the evaluation of nonlinear distortion effects in inter-satellite microwave photonics links using a dual-drive Mach-Zehnder modulator (DD-MZM) under dual-tone modulation. The model results in simple exact expressions for any harmonic and inter-modulation components at the output of the detector which can be applied to almost all operating conditions of DD-MZM, providing useful information to select system parameters for the performance optimization. Two special cases, double sideband and single sideband modulation, are studied in detail. Numerical results for the third-order inter-modulation distortion ratio (IMR3) are well matched with the analysis ones.     

Nylon and nylon blend nanotubes and nanorods

Nylon nanorods and nanotubes (200 nm diameter) were fabricated by the membrane wetting technique (solvent and melt wetting) from a range of nylons (6; 6,6; 6,9; 6,10; 6,12; 11; 12, 6(3)T) and nylon blended with different dyes (Nylon Cast Blue, Nylon 6/6 Black) or with molybdenum disulfide (Nylon cast MDS). The 65-μm long nylon nanotubes and nanorods were characterized by scanning electron microscopy. The nanoscale nylon 6,6 served as an effective high surface area alternative to a nylon membrane as a solid support in a chemiluminescent assay for nylon-bound biotinylated nucleic acids based on streptavidin- alkaline phosphatase and chemiluminescent detection of the bound alkaline phosphatase label with the dioxetane substrate, CDP-Star. Layer-by-layer deposition of the cationic polymer (Sapphire-II™; Tropix) onto the nylon 6,6 nanostructures prior to UV-cross-linking with biotinylated DNA resulted in further enhancement of binding and detection of biotinylated DNA.     

Effects of cryopreservation on developmental competency, cytological and molecular stability of citrus callus

Cell suspensions of twelve citrus genotypes were successfully cryopreserved by vitrification. All genotypes survived cryopreservation with >90% viability and the surviving cells of some genotypes regenerated somatic embryos better than the controls. Single-cell sibling lines of cultivar Newhall were used for cytological and molecular examination. It was found that the ploidy constitution remained genetically stable and that no DNA sequence variation was detected by randomly amplified polymorphic DNA (RAPD) assay after cryopreservation. In addition, the methylation sensitive amplified polymorphism (MSAP) assay indicated that cryopreservation caused a significant change in DNA methylation status.     

DNA Binding Property and Antitumor Evaluation of Xanthone with Dimethylamine Side Chain

In this work, a xanthone derivative was obtained by cationic modification of the free hydroxyl group of xanthone with dimethylamine group of high pKa value. The interactions of xanthones with DNA were investigated by spectroscopic methods, electrophoretic migration assay and polymerase chain reaction test. Results indicate that xanthones can intercalate into the DNA base pairs by the hydrophobic plane and the xanthone with dimethylamine side chain may also bind the DNA phosphate framework by the basic amine alkyl chain, thus showing a better DNA binding ability than the xanthone. Furthermore, inhibition on tumor cells (ECA109, SGC7901, GLC-82) proliferation of xanthones were evaluated by MTT method. Analysis results show that the xanthone with dimethylamine side chain exhibits more effective inhibition activity against three cancer cells than the xanthone. The effects on the inhibition of tumor cells in vitro agree with the studies of DNA binding. It means that the amine alkyl chain would play an important role in its antitumor activity and DNA binding property.     

Das Institut für Metallphysik und Reinstmetalle,Dresden, der Deutschen Akademie der Wissenschaften zu Berlin

Ein Beitrag des Instituts für Metallphysik und Reinstmetalle (IMR) im Jubiläumsheft zum lOjährigen Bestehen der Kernforschung und Kerntechnik in der DDR dürfte von allgemeinem Interesse sein, da die Arbeiten des Instituts zum Teil in enger Beziehung zur Kernforschung stehen. Das trifft besonders zu für die Arbeiten auf dem Gebiet der Strahlenchemie.     

On the mechanism of effective chemical reactions with turbulent mixing of reactants and finite rate of molecular reactions

A generalization of the theory of chemical transformation processes under turbulent mixing of reactants and arbitrary values of the rate of molecular reactions is presented that was previously developed for the variant of an instantaneous reaction [13]. The use of the features of instantaneous reactions when considering the general case, namely, the introduction of the concept of effective reaction for the reactant volumes and writing a closing conservation equation for these volumes, became possible due to the partition of the whole amount of reactants into “active” and “passive” classes; the reactants of the first class are not mixed and react by the mechanism of instantaneous reactions, while the reactants of the second class approach each other only through molecular diffusion, and therefore their contribution to the reaction process can be neglected. The physical mechanism of reaction for the limit regime of an ideal mixing reactor (IMR) is revealed and described. Although formally the reaction rate in this regime depends on the concentration of passive fractions of the reactants, according to the theory presented, the true (hidden) mechanism of the reaction is associated only with the reaction of the active fractions of the reactants with vanishingly small concentration in the volume of the reactor. It is shown that the rate constant of fast chemical reactions can be evaluated when the mixing intensity of reactants is much less than that needed to reach the mixing conditions in an IMR.     

重离子对人胃癌细胞DNA错配修复基因MSH2 表达的影响

采用高传能线密度(LET) 重离子辐照人胃癌SGC7901 细胞,应用流式细胞技术、蛋白质印迹法(Western blot) 及反转录聚合酶链式反应(RT-PCR) 观察重离子诱导人胃癌SGC7901 细胞周期、凋亡和MSH2 表达状况。结果表明: 与对照组相比,SGC7901 细胞在辐射后72 h G2/M 期所占细胞比率(33.26±0.08) 和凋亡率(24.16±0.64) 均达到峰值,且呈时间依赖性增加;经重离子照射后,DNA错配修复基因MSH2 mRNA 和蛋白表达水平在6 h 最高。结果提示:重离子在体外诱导SGC7901 细胞周期阻滞和凋亡,且具有显著的时间依赖性效应;重离子在一定剂量和时间下,诱导了SGC7901 细胞MSH2 基因表达。DNA错配修复基因MSH2 可能参与了重离子辐照诱导胃癌细胞DNA损伤的修复应答。Human gastric cancer cell SGC7901 were irradiated with high linear energy transfer (LET) carbon ion. Apoptotic cells after irradiation were analyzed by flow cytometry and expression of MSH2 genes in the irradiated cells was detected by western blot and RT-PCR assay. Compared with the control group, we found that the number of G2/M (33.26±0.08) or apoptosis (24.16±0.64) of SGC7901 cells reached a maximum after irradiation at 72 h in a dose dependent manner. And heavy ion irradiation efficiently up-regulated the expression of MSH2 gene at 4.0 Gy after being irradiated 6 h. These results imply that heavy ion beam could induce cell apoptosis and cell cycle arrest in time-dependent manners. Furthermore, expression of MSH2 genes activated by carbon ion irradiation suggests that DNA mismatch repair gene MSH2 might be involved in DNA repair pathways.     

Development of a Fluorescent Enzyme-Linked DNA Aptamer-Magnetic Bead Sandwich Assay and Portable Fluorometer for Sensitive and Rapid Leishmania Detection in Sandflies

A fluorescent peroxidase-linked DNA aptamer-magnetic bead sandwich assay is described which detects as little as 100 ng of soluble protein extracted from Leishmania major promastigotes with a high molarity chaotropic salt. Lessons learned during development of the assay are described and elucidate the pros and cons of using fluorescent dyes or nanoparticles and quantum dots versus a more consistent peroxidase-linked Amplex Ultra Red (AUR; similar to resazurin) fluorescence version of the assay. While all versions of the assays were highly sensitive, the AUR-based version exhibited lower variability between tests. We hypothesize that the AUR version of this assay is more consistent, especially at low analyte levels, because the fluorescent product of AUR is liberated into bulk solution and readily detectable while fluorophores attached to the reporter aptamer might occasionally be hidden behind magnetic beads near the detection limit. Conversely, fluorophores could be quenched by nearby beads or other proximal fluorophores on the high end of analyte concentration, if packed into a small area after magnetic collection when an enzyme-linked system is not used. A highly portable and rechargeable battery-operated fluorometer with on board computer and color touchscreen is also described which can be used for rapid (<1 h) and sensitive detection of Leishmania promastigote protein extracts (～100 ng per sample) in buffer or sandfly homogenates for mapping of L. major parasite geographic distributions in wild sandfly populations.     

Study on the Interaction of the CpG Alternating DNA with CdTe Quantum Dots

A novel sensitive method for detection of DNA methylation was developed with thioglycollic acid (TGA)-capped CdTe quantum dots (QDs) as fluorescence probes. Recognition of methylated DNA sites would be useful strategy due to the important roles of methylation in disease occurrence and developmental processes. DNA methylation occurs most often at cytosine-guanine sites (CpG dinucleotides) of gene promoters. The QDs significantly interacted with hybridized unmethylated and methylated DNA. The interaction of CpG rich methylated and unmethylated DNA hybrid with quantum dots as an optical probe has been investigated by fluorescence spectroscopy and electrophoresis assay. The fluorescence intensity of QDs was highly dependent to unmethylated and methylated DNA. Specific site of CpG islands of Adenomatous polyposis coli (APC), a well-studied tumor suppressor gene, was used as the detection target. Under optimum conditions, upon the addition of unmethylated dsDNA, the fluorescence intensity increased in linear range from 1.0?×?10^??10 to 1.0?×?10^??6M with detection limit of 6.2?×?10^??11 M and on the other hand, the intensity of QDs showed no changes with addition of methylated dsDNA. We also demonstrated that the unmethylated and methylated DNA and QDs complexes showed different mobility in electrophoresis assay. This easy and reliable method could distinguish between methylated and unmethylated DNA sequences.     

吖啶黄反应光度法测定脱氧核糖核酸含量的研究

以微量脱氧核糖核酸与吖啶黄的反应为基础,优化了实验条件。基于DNA与吖啶黄最大吸收波长444nm的吸光度有线性降低的关系,建立了测定微量DNA的新方法。在0～8.0μg.mL-1浓度呈现良好的线性关系,相关系数r为0.9998,检出限为0.12μg.mL-1。初步探讨了反应机理,认为DNA与吖啶黄通过嵌入作用生成了加合物。     

Improvements in the Sensitivity of Time Resolved Fluorescence Energy Transfer Assays

The effect on fluorescence resonance energy transfer (FRET) of multiple labelling of DNA oligonucleotides with donor lanthanide chelate and acceptor CyDye fluors has been investigated. It is shown that using a multiple donor lanthanide chelate with a single acceptor Cy or Cy5 can increase sensitivity and fluorescence output. The enhanced FRET observed in the multiple donor label system has been utilised in two different DNA based assay formats to demonstrate the advantages over a steady state fluorescence assay and a radiometric assay.     

Formation of Direct and Enzymatic DNA Double-Strand Breaks in the Presence of Repair Inhibitors after Exposure to Radiations of Different Quality

With the use of the DNA comet assay and immunocytochemical staining, regularities have been studied in the induction and repair of DNA double-strand breaks(DSBs) in human cells after exposure to ⁶⁰Co γ-rays and accelerated heavy ions with different linear energy transfer (LET) in the presence of the DNA repair inhibitors cytosine arabinoside and hydroxyurea. It is shown that for heavy ions the agents’ modifying effect decreases with increasing particles’ LET. The approach involving DNA synthesis inhibitors used in this study allows an estimation of the proportion of enzymatic DNA DSBs in total DSB yield after exposure to ionizing radiations of different quality.