Early appearance of 2, 3-butanediol in acute myocardial infarction. A new marker for ischaemia?

In 28 patients with acute myocardial infarction, the release pattern of 2, 3-butanediol (BD), a product of intermediary metabolism, and creatine kinase activity (CK) in blood were compared. Whereas CKat entry was low in all patients, the BD level was elevated in 18 (64%). However, BD returned to normal levels during the next 24 h whereas CK increased. The BD level at entry did not allow differentiation between patients with transmural or non-transmural infarction; it was independent of clinical findings and biochemical parameters. We suggest that, in patients with acute myocardial infarction, elevated levels of BD originates from myo-cardial metabolism. Whether it reflects ongoing ischaemia or reperfusion of the infarcted area remains unresolve

Direct fluorescent labelling of clones by DOP PCR

<p>Abstract</p> <p>Background</p> <p>Array Comparative Genomic Hybridisation (array CGH) is a powerful technique for the analysis of constitutional chromosomal anomalies. Chromosomal duplications or deletions detected by array CGH need subsequently to be validated by other methods. One method of validation is Fluorescence <it>in situ </it>Hybridisation (FISH). Traditionally, fluorophores or hapten labelling is performed by nick translation or random prime labelling of purified Bacterial Artificial Chromosome (BAC) products. However, since the array targets have been generated from Degenerate Oligonucleotide Primed (DOP) amplified BAC clones, we aimed to use these DOP amplified BAC clones as the basis of an automated FISH labelling protocol. Unfortunately, labelling of DOP amplified BAC clones by traditional labelling methods resulted in high levels of background.</p> <p>Results</p> <p>We designed an improved labelling method, by means of degenerate oligonucleotides that resulted in optimal FISH probes with low background.</p> <p>Conclusion</p> <p>We generated an improved labelling method for FISH which enables the rapid generation of FISH probes without the need for isolating BAC DNA. We labelled about 900 clones with this method with a success rate of 97%.</p

2,3-butanediol in experimental myocardial ischaemia in pigs

To investigate the role of 2,3-butanediol in myocardial ischaemia we analysed this compound in pig's myocardium and blood. Ischaemia was induced by ligation of a coronary artery. In the first study we found significantly higher levels of 2,3-butanediol in the homogenate of ischaemic myocardium than in non-ischaemic myocardium. The lactate concentration was also significantly elevated. In the second study, where ischaemia was similarly induced, and where reperfusion was achieved by re-opening the ligated coronary artery after 20 min, 2,3-butanediol in peripheral blood was found to increase significantly. In the pigs in which the coronary artery was not re-opened, the 2,3-butanediol level in peripheral blood was unchanged. We conclude that in pigs' anaerobic myocardia accumulation of 2,3-butanediol occurs; if the myocardium is reperfused this metabolite also appears in the bloo

Array Formatting of the Heat-Transfer Method (HTM) for the Detection of Small Organic Molecules by Molecularly Imprinted Polymers

In this work we present the first steps towards a molecularly imprinted polymer (MIP)-based biomimetic sensor array for the detection of small organic molecules via the heat-transfer method (HTM). HTM relies on the change in thermal resistance upon binding of the target molecule to the MIP-type receptor. A flow-through sensor cell was developed, which is segmented into four quadrants with a volume of 2.5 μL each, allowing four measurements to be done simultaneously on a single substrate. Verification measurements were conducted, in which all quadrants received a uniform treatment and all four channels exhibited a similar response. Subsequently, measurements were performed in quadrants, which were functionalized with different MIP particles. Each of these quadrants was exposed to the same buffer solution, spiked with different molecules, according to the MIP under analysis. With the flow cell design we could discriminate between similar small organic molecules and observed no significant cross-selectivity. Therefore, the MIP array sensor platform with HTM as a readout technique, has the potential to become a low-cost analysis tool for bioanalytical applications

Defining a Novel Role for the Coxsackievirus and Adenovirus Receptor in Human Adenovirus Serotype 5 Transduction In Vitro in the Presence of Mouse Serum

Human adenoviral serotype 5 (HAdV-5) vectors have predominantly hepatic tropism when delivered intravascularly, resulting in immune activation and toxicity. Coagulation FX binding to HAdV-5 mediates liver transduction and provides protection from virion neutralisation in mice. FX is dispensable for liver transduction in mice lacking IgM antibodies or complement, suggesting alternative transduction pathways exist. To identify novel factor(s) mediating HAdV-5 FX-independent entry, we investigated HAdV-5 transduction in vitro in the presence of serum from immunocompetent C57BL/6 or immunocompromised mice lacking IgM antibodies (Rag 2-/- and NSG). Sera from all three mouse strains enhanced HAdV-5 transduction of A549 cells. While inhibition of HAdV-5:FX interaction with X-bp inhibited transduction in the presence of C57BL/6 serum, it had negligible effect on the enhanced transduction observed in the presence of Rag 2-/- or NSG serum. Rag 2-/- serum also enhanced transduction of the FX-binding deficient AdT*. Interestingly, Rag 2-/- serum enhanced HAdV-5 transduction in a FX-independent manner in CHO-CAR and SKOV3-CAR cells. Additionally, blockade of CAR with soluble HAdV-5 fiber knob inhibited mouse serum-enhanced transduction in A549 cells, suggesting a potential role for CAR. Transduction of HAdV-5 KO1 and HAdV-5/F35 (CAR-binding deficient) in the presence of Rag 2-/- serum was equivalent to that of HAdV-5, indicating that direct interaction between HAdV-5 and CAR is not required. These data suggest that FX may protect HAdV-5 from neutralization but has minimal contribution to HAdV-5 transduction in the presence of immunocompromised mouse serum. Alternatively, transduction occurs via an unidentified mouse serum protein capable of bridging HAdV-5 to CAR

Label-free protein detection based on the heat-transfer method-a case study with the peanut allergen Ara h 1 and aptamer-based synthetic receptors

© 2015 American Chemical Society. Aptamers are an emerging class of molecules that, because of the development of the systematic evolution of ligands by exponential enrichment (SELEX) process, can recognize virtually every target ranging from ions, to proteins, and even whole cells. Although there are many techniques capable of detecting template molecules with aptamer-based systems with high specificity and selectivity, they lack the possibility of integrating them into a compact and portable biosensor setup. Therefore, we will present the heat-transfer method (HTM) as an interesting alternative because this offers detection in a fast and low-cost manner and has the possibility of performing experiments with a fully integrated device. This concept has been demonstrated for a variety of applications including DNA mutation analysis and screening of cancer cells. To the best our knowledge, this is the first report on HTM-based detection of proteins, in this case specifically with aptamer-type receptors. For proof-of-principle purposes, measurements will be performed with the peanut allergen Ara h 1 and results indicate detection limits in the lower nanomolar regime in buffer liquid. As a first proof-of-application, spiked Ara h 1 solutions will be studied in a food matrix of dissolved peanut butter. Reference experiments with the quartz-crystal microbalance will allow for an estimate of the areal density of aptamer molecules on the sensor-chip surface

P5.8 - Light-stimulated hydrogel actuators with incorporated graphene oxide for microfluidic applications

Graphene oxide (GO) nanoparticles, which are able to convert light energy into heat, were incorporated into temperature-sensitive Poly-N-isopropylacrylamide (P-NIPAAm) hydrogels. In contrast to pure P-NIPAAm hydrogels, the new polymers with GO showed significant sensitivity towards light as a stimulus. The photothermal conversion and heating process of hydrogels under illumination were investigated by IRthermography. Furthermore, first experiments for a possible application as light-driven microvalves within lab-on-chip systems were performed