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Nanotechnology and its broad range of applications are becoming more and more important for modern society. It is said to be one of the key technologies of the current century. This article shows ways of implementing nanotechnology in schools. The presented syntheses of gold and silver nanoparticles were carried out in a miniaturized lab and adapted for use in class. They are appropriate to be carried out by the pupils themselves in school or in science centers, in brief or in longer projects. Furthermore, a student-centered teaching concept for a complete nanotechnology unit focused on key qualifications as claimed by the national curricula is presented. Students can learn about scientific work methods and get an insight view of scientific daily routine. [ABSTRACT FROM AUTHOR]

This work describes the optimization and adoption of an assay system for the Human Immunodeficiency Virus (HIV)-protease, whose inhibition plays a central role in HIV therapy. The HIV-protease, which is an essential enzyme during viral maturation, has a specific cleavage site of eight amino acid residues (SQNY*PIV). Adding two amino acid residues at the N-terminus and enclosing the resulting sequence by a dye-labelled lysine residue and a tryptophan residue leads to the substrate (K(dye)CGSQNY*PIVW) in which the fluorescence of the fluorophore is efficiently quenched by the intrinsic tryptophan due to a photoinduced electron transfer reaction. After cleavage of the substrate by the target enzyme, the dye and the tryptophan residue are separated, effecting a significant increase in fluorescence intensity. Measuring the fluorescence versus time enables an online-monitoring of the enzyme activity. With this method, a HIV-PR concentration of 10-9 M is detectable within minutes, which is comparable with commercially available assays using doubly labelled substrates based on a fluorescence resonance energy transfer. We were able to further increase the sensitivity to the subnanomolar range by using confocal single-molecule spectroscopy. [ABSTRACT FROM AUTHOR]

In diesem Jahr wird bundesweit der 125. Entdeckungstag der Röntgenstrahlen und der 175. Geburtstag ihres Entdeckers Wilhelm Conrad Röntgen gefeiert. Für das Röntgenjahr 2020 haben sich das Deutsche Röntgen‐Museum (DRM) in Remscheid und die Reiss‐Engelhorn‐Museen (rem) in Mannheim ein X‐perimente Mobil konzipiert und mit zahlreichen Experimenten und Gerätesets ausgestattet, das Schulen und Bildungseinrichtungen anfährt. [ABSTRACT FROM AUTHOR]

This paper presents new self-quenching DNA probes that exploit the efficient fluorescence quenching by the formation of dye dimers. The probes consist of a hairpin-structured oligonucleotide that is labeled with two identical fluorescence dyes that are able to form non-fluorescent H-type dimers while the hairpin is closed. We used the oxazine derivative MR 121 that has a sufficient dimerization tendency and can be excited by a pulsed diode laser emitting at 635  nm. Upon hybridization to the target DNA, the dyes are separated and a 12-fold increase of the fluorescence intensity occurs. The probe was used for the specific detection of Mycobacterium xenopi in a model system. Specific target DNA and a control target, differing by six nucleotides were amplified by polymerase chain reaction (PCR). A confocal fluorescence microscope was used to observe the fluorescence bursts of individual probe molecules in the presence of target PCR product and controls. By experiment and by the respective simulation we demonstrated that the secondary structure of the target DNA hinders the hybridization to the DNA probe at room temperature. Based on these data a successful hybridization procedure was developed and allowing the detection of nanomolar concentrations of Mycobacterium xenopi specific target at room temperature, using single-molecule detection techniques. [ABSTRACT FROM AUTHOR]

In recent years, the interest in assaying exopeptidases has become increasingly important because they are significantly involved in many diseases like cancer. To date, no generally applicable fluorescence-based detection method has been developed because commercially available doubly-labeled substrates are not always digested by exopeptidases. In this article we present a new method for the sensitive detection of exopeptidases based on fluorescently-labeled substrates containing only one fluorophore that is efficiently quenched by an adjacent tryptophan residue via photoinduced electron transfer. Because of their well-known properties we chose carboxypeptidase A (CPA) as a model system. The self-quenching probes were used in homogeneous solution as well as on cross-linked PEG-coated surfaces in combination with single-molecule imaging techniques. However, even with standard fluorescence spectrometers we achieved sensitivity below the picomolar range. [ABSTRACT FROM AUTHOR]

In this article, we describe a new method for single nucleotide polymorphism analysis using the displacement reaction of the DNA in a PNA–DNA double helix by the target DNA. Thereby, the probe consists of a TMR-labeled PNA and a Cy5-labeled DNA forming a FRET system. Due to the displacement of the labeled DNA strand by the target DNA, the FRET is invalidated and the fluorescence of the donor dye (TMR) increases. Investigations of the exchange reaction show that increasing salt concentration and temperature accelerate the exchange rate. [ABSTRACT FROM AUTHOR]

Based on a single-molecule sensitive fluorescence-linked immunosorbent assay, an analytical platform for the detection of lipoarabinomannan (LAM), a lipopolysaccharide marker of tuberculosis, was established that is about 3 orders of magnitude more sensitive than comparable current ELISA assays. No amplification step was required. Also, no particular sample preparation had to be done. Since individual binding events are detected, true quantification was possible simply by counting individual signals. Utilizing a total internal reflection configuration, unprocessed biological samples (human urine and plasma) to which LAM was added could be analyzed without the requirement of sample purification or washing steps during analysis. Samples containing about 600 antigen molecules per microliter produced a distinct signal. The methodology developed can be employed for any set of target molecules for which appropriate antibodies exist.

Steady-state and time-resolved fluorescence measurements were performed to elucidate the fluorescence quenching of oxazine, rhodamine, carbocyanine, and bora-diaza-indacene dyes by amino acids. Among the natural amino acids, tryptophan exhibits the most pronounced quenching efficiency. Especially, the red-absorbing dyes ATTO 655, ATTO 680, and the oxazine derivative MR 121 are strongly quenched almost exclusively by tryptophan due to the formation of weak or nonfluorescent ground-state complexes with association constants, K(ass.), ranging from 96 to 206 M(-1). Rhodamine, fluorescein, and bora-diaza-indacene derivatives that absorb at shorter wavelengths are also quenched substantially by tyrosine residues. The quenching of carbocyanine dyes, such as Cy5, and Alexa 647 by amino acids can be almost neglected. While quenching of ATTO 655, ATTO 680, and the oxazine derivative MR121 by tryptophan is dominated by static quenching, dynamic quenching is more efficient for the two bora-diaza-indacene dyes Bodipy-FL and Bodipy630/650. Labeling of the dyes to tryptophan, tryptophan-containing peptides, and proteins (streptavidin) demonstrates that knowledge of these fluorescence quenching processes is crucial for the development of fluorescence-based diagnostic assays. Changes in the fluorescence quantum yield of dye-labeled peptides and proteins might be used advantageously for the quantification of proteases and specific binding partners.