Improvement of bioanalytical parameters through automation: suitability of a hand-like robotic system.

Commercial automation systems for small- and medium-sized laboratories, including research environments, are often complex to use. For liquid handling systems (LHS), development is required not only for the robot's movements but also for adapting the bioanalytical method to the automated system. This study investigates whether a more human-like automation strategy-using a robotic system (RS)-is more suitable for research laboratories than a professional automation approach utilizing a commercial automated LHS.

Can I benefit from laboratory automation? A decision aid for the successful introduction of laboratory automation.

The large volumes of samples to be analysed every day would be impossible to manage without laboratory automation. As laboratory procedures have progressed, so have the tasks of laboratory personnel. With this feature article, we would like to provide (bio)chemical practitioners with little or no knowledge of laboratory automation with a guide to help them decide whether to implement laboratory automation and find a suitable system.

An expandable voice user interface as lab assistant based on an improved version of Google's speech recognition.

Voice assistants are potentially helpful when working in a scientific laboratory. A big challenge is the extremely specific use of language in every laboratory. As with any voice assistant, another concern is data security.

Establishment of low-cost laboratory automation processes using AutoIt and 4-axis robots.

In most small laboratories, many processes are not yet automated because existing laboratory automation solutions are usually expensive and inflexible to use. Examples of this are autosamplers that are only compatible with one specific laboratory instrument or larger liquid handling stations that are expensive and usually self-contained. A flexible and inexpensive way to automate laboratory processes would be to automate existing laboratory equipment with the help of suitable robotic arms.

Proline-Rich Antimicrobial Peptides in Medicinal Maggots of Interact With Bacterial DnaK But Do Not Inhibit Protein Synthesis.

In the search for new antibiotics to combat multidrug-resistant microbes, insects offer a rich source of novel anti-infectives, including a remarkably diverse array of antimicrobial peptides (AMPs) with broad activity against a wide range of species. Larvae of the common green bottle fly are used for maggot debridement therapy, and their effectiveness in part reflects the large panel of AMPs they secrete into the wound. To investigate the activity of these peptides in more detail, we selected two structurally different proline rich peptides (Lser-PRP2 and Lser-PRP3) in addition to the α-helical peptide Lser-stomoxyn.

Rational design of a low-affinity peptide for the detection of cystatin C in a fast homogeneous immunoassay.

Immunoassays play an essential role in current research and diagnostics resulting in a variety of detection principles. Thereby, homogeneous assays are often used for a fast signal response as demanded for example in point-of-care diagnostics. These systems often rely on a competitive assay design where the sample analyte and the corresponding dye-labeled substance are competing for binding sites on an antibody present in limited amounts.

Impaired APP activity and altered Tau splicing in embryonic stem cell-derived astrocytes obtained from an APPsw transgenic minipig.

Animal models of familial juvenile onset of Alzheimer's disease (AD) often fail to produce diverse pathological features of the disease by modification of single gene mutations that are responsible for the disease. They can hence be poor models for testing and development of novel drugs. Here, we analyze in vitro-produced stem cells and their derivatives from a large mammalian model of the disease created by overexpression of a single mutant human gene (APPsw).

Insect antimicrobial peptides show potentiating functional interactions against Gram-negative bacteria.

Antimicrobial peptides (AMPs) and proteins are important components of innate immunity against pathogens in insects. The production of AMPs is costly owing to resource-based trade-offs, and strategies maximizing the efficacy of AMPs at low concentrations are therefore likely to be advantageous. Here, we show the potentiating functional interaction of co-occurring insect AMPs (the bumblebee linear peptides hymenoptaecin and abaecin) resulting in more potent antimicrobial effects at low concentrations.

Protein surface charge of trypsinogen changes its activation pattern.

Background: Trypsinogen is the inactive precursor of trypsin, a serine protease that cleaves proteins and peptides after arginine and lysine residues. In this study, human trypsinogen was used as a model protein to study the influence of electrostatic forces on protein-protein interactions. Trypsinogen is active only after its eight-amino-acid-long activation peptide has been cleaved off by another protease, enteropeptidase.

One-step assay for the quantification of T4 DNA ligase.

As one of the most commonly used enzyme in molecular biology, the T4 DNA ligase presents an important tool for the manipulation of DNA. T4 DNA ligase activity measurements are based on the use of radioactivity or rather labor-intense procedures including gel-based analysis. We therefore established a homogeneous T4 DNA ligase assay utilizing a specifically designed fluorescein- and dark quencher-labeled DNA molecule.

A homogeneous assay principle for universal substrate quantification via hydrogen peroxide producing enzymes.

H2O2 is a widely occurring molecule which is also a byproduct of a number of enzymatic reactions. It can therefore be used to quantify the corresponding enzymatic substrates. In this study, the time-resolved fluorescence emission of a previously described complex consisting of phthalic acid and terbium (III) ions (PATb) is used for H2O2 detection.

Improvement of an antibody-enzyme coupling yield by enzyme surface supercharging.

Background: Protein cross-coupling reactions demand high yields, especially if the products are intended for bioanalytics, like enzyme-linked immunosorbent assays. Amongst other factors, the coupling yield depends on the concentration of the proteins being used for coupling. Protein supercharging of enzymes can increase the solubility dramatically, which could promote enzyme-antibody coupling reactions.

His-tag protein monitoring by a fast mix-and-measure immunoassay.

Here, we present a fast mix-and-measure immunoassay for the specific semiquantitative detection of His-tagged proteins, for example in E. coli cell lysate. The assay is based on Förster resonance energy transfer (FRET) between a lanthanide dye-labeled low-affinity His-peptide and an acceptor-labeled anti-His-tag antibody.

A luminescence-based probe for sensitive detection of hydrogen peroxide in seconds.

Here, we present a fast and simple hydrogen peroxide assay that is based on time-resolved fluorescence. The emission intensity of a complex consisting of terbium ions (Tb(3+)) and phthalic acid (PA) in HEPES buffer is quenched in the presence of H2O2 and this quenching is concentration-dependent. The novel PATb assay detects hydrogen peroxide at a pH range from 7.

Aggregation behavior of chemically synthesized, full-length huntingtin exon1.

Repeat length disease thresholds vary among the 10 expanded polyglutamine (polyQ) repeat diseases, from about 20 to about 50 glutamine residues. The unique amino acid sequences flanking the polyQ segment are thought to contribute to these repeat length thresholds. The specific portions of the flanking sequences that modulate polyQ properties are not always clear, however.

Highly efficient förster resonance energy transfer in a fast, serum-compatible immunoassay.

Highly efficient FRET leads to important enhancements for homogeneous immunoassays. By using the novel phosphorescent dye EuLH and BHQ-10 as a donor-acceptor pair, the FRET efficiency increases to >99.5 %, leading to significantly improved signal-to-background ratio, precision and linear range.

Crystal structure of a supercharged variant of the human enteropeptidase light chain.

The highly specific serine protease human enteropeptidase light chain cleaves the Asp4Lys recognition sequence and represents an interesting enzyme for biotechnological applications. The human enzyme shows 10 times faster kinetics compared to other animal sources but low solubility under low salt conditions, which hampers protein production and crystallization. Therefore, a supercharged variant (N6D/G21D/G22D/N142D/K210E/C112S) with increased solubility was used for crystallization.

Highly adaptable and sensitive protease assay based on fluorescence resonance energy transfer.

Proteases are widely used in analytical sciences and play a central role in several widespread diseases. Thus, there is an immense need for highly adaptable and sensitive assays for the detection and monitoring of various proteolytic enzymes. We established a simple protease fluorescence resonance energy transfer (pro-FRET) assay for the determination of protease activities, which could in principle be adapted for the detection of all proteases.

Homogeneous fluorescence-based immunoassay detects antigens within 90 seconds.

Homogeneous immunoassays are prevalent tools for the detection of antigens. The major advantage over heterogeneous immunoassays is the absence of numerous incubation and washing steps, reducing the assay time and allowing rapid on-site detection of antigens (e.g.

Surface supercharged human enteropeptidase light chain shows improved solubility and refolding yield.

Enteropeptidase is a serine protease used in different biotechnological applications. For many applications the smaller light chain can be used to avoid the expression of the rather large holoenzyme. Recombinant human enteropeptidase light chain (hEPL) shows high activity but low solubility and refolding yields, currently limiting its use in biotechnological applications.

Synthesis of pathological and nonpathological human exon 1 huntingtin.

Huntington's disease (HD) is a neurodegenerative disorder that affects approximately 1 in 10 000 individuals. The underlying gene mutation was identified as a CAG-triplet repeat expansion in the gene huntingtin. The CAG sequence codes for glutamine, and in HD, an expansion of the polyglutamine (poly-Q) stretch above 35 glutamine residues results in pathogenicity.

Highly sensitive protein detection based on lanthanide chelates with antenna ligands providing a linear range of five orders of magnitude.

Protein detection is an important task for pharmaceutical and clinical research today. Numerous protein staining techniques exist but are limited regarding their sensitivity and often narrow linear quantification ranges. To the best of our knowledge, this is the first description of a novel class of lanthanide chelatators, which absorb in the lower energy region at 360 nm.

Identification of proteins involved in neural progenitor cell targeting of gliomas.

Background: Glioblastoma are highly aggressive tumors with an average survival time of 12 months with currently available treatment. We have previously shown that specific embryonic neural progenitor cells (NPC) have the potential to target glioma growth in the CNS of rats. The neural progenitor cell treatment can cure approximately 40% of the animals with malignant gliomas with no trace of a tumor burden 6 months after finishing the experiment.

Instructive cross-talk between neural progenitor cells and gliomas.

Gliomas are the most common primary brain tumors and offer a poor prognosis in patients because of their infiltrative and treatment-resistant nature. The median survival time after diagnosis is approximately 11-12 months. There is a strong need for novel treatment modalities in targeting gliomas, and recent advances use neural progenitor cells as delivery systems for different therapeutic strategies.

Down-regulation of muscarinic acetylcholine receptor M2 adversely affects the expression of Alzheimer's disease-relevant genes and proteins.

Beta-amyloid peptides play a major role in the pathogenesis of Alzheimer's disease (AD). Therefore, preventing beta-amyloid formation by inhibition of the beta site amyloid precursor protein-cleaving enzyme (BACE) 1 is considered as a potential strategy to treat AD. Cholinergic mechanisms have been shown to control amyloid precursor protein processing and the number of muscarinic M2-acetylcholine receptors is decreased in brain regions of patients with AD enriched with senile plaques.