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With the concomitant increase of blood transfusions and safety rules, there is a growing need to integrate high-throughput and multiparametric assays within blood qualification centers. Using a robust and automated solution, we describe a new method for extended blood group genotyping (HiFi-Blood 96) bringing together the throughput possibilities of complete automation and the microarray multiplexed analysis potential. Our approach provides a useful resource for upgrading blood qualification center facilities. A set of six single-nucleotide polymorphisms (SNPs) associated with clinically important blood group antigens (Kell, Kidd, Duffy, and MNS systems) were selected and the corresponding genotyping assays developed. A panel of 293 blood samples was used to validate the approach. The resulting genotypes were compared to phenotypes previously determined by standard serologic techniques, and excellent correlations were found for five SNPs out of six. For the Ken, Kidd, Duffy, and MNS3/MNS4 systems, high matching percentages of 100%, 98.9%, 97.7%, and 97.4% were obtained, respectively, whereas a concordance percentage of 83.3% only was attained for the MNS1/MNS2 polymorphism. 10.1021/ac101008d

The effect of the ionic liquid 1-ethyl-3-methylimidazolium ethylsulfate ([Emim][EtS[O.sub.4]]) on the copper-catalyzed luminol chemiluminescence (CL) is reported. A drastic light emission enhancement is observed, related to a strong interaction between [Cu.sup.2+] and the imidazolium ring. In these conditions, the CL reaction was able to produce light efficiently at pH as low as 6.5 (amplification factor: [Intensity.sup.+IL]/[Intensity.sup.-IL] = 2900). Interesting effects of [Emim][EtS[O.sub.4]] on the enzyme glucose oxidase activity were also evidenced, and advantages were taken from this enhancement to perform sensitive chemiluminescent glucose detection (LOD = 4 [micro]M) at 10.1021/ac9026725

Byline: Agnes P. Girard-Egrot, Christophe A. Marquette, Loic J. Blum Biological membranes constitute a source of inspiration for making supramolecular assemblies which can be used in the design of biomimetic sensors. At the same time, the concept of using biomolecules as an elementary structure to develop self-assembled entities has received considerable attention. More particularly, the ability of amphiphilic molecules like lipids, to spontaneously organise into bilayers, is suitable to achieve biomimetic membrane models. The potential of two-dimensional molecular self-assemblies is clearly illustrated by Langmuir monolayers of lipids formed at an air/water interface, which can be used as models to acquire knowledge about the molecular recognition process occurring in biological membranes. Langmuir-Blodgett (LB) technology, based on the transfer of this interfacial monomolecular film onto a solid support, allows building up lamellar lipid stack, with an accurate control of thickness and molecular organisation. This technique offers the possibility to prepare ultrathin layers suitable for biomolecule immobilisation. We are presenting herein an overview of work performed in our group that sheds light on the formation of biomimetic LB membranes associating protein in a well-defined orientation. Two points will be addressed: investigations of protein/lipid interactions using lipid monolayers as membrane models and biosensing applications. The objectives are to highlight advantages of interfacial Langmuir monolayers and supported Langmuir-Blodgett films to investigate molecular interactions between biomolecules and lipid membrane components or to elaborate biomimetic membranes as sensing layers, respectively. The present paper also draws a general picture of non-conventional methods for biomolecule immobilisation and their applications for biochip developments. The technologies presented are based either on original solid supports or on innovative immobilisation processes. First, 'Macromolecules to PDMS transfer' technique relying on the direct entrapment of macromolecules spots during PDMS polymerisation is proposed as an alternative for the easy and simple PDMS surface modification. Then, the electro-addressing of biomolecule-aryl diazonium adducts at the surface of conducting biochips will be presented and shown to be an interesting alternative to immobilisation processes based on surface functionalisation.

The progress in the development of DNA biosensors and microarrays which could be applied to probe immobilization and hybridization detection are reviewed.

Screen-printed (SP) graphite electrode microarrays are modified with electro-addressed diazonium cations, leading to an innovative approach to electro-address immobilization of proteins. Biomolecules are first modified with the aniline derivative 4-carboxymethylaniline (CMA) and then successfully electrodeposited, without parasitic reduction of the protein, at the graphite surface through an optimized protocol compatible with biomolecules like rabbit and human antibodies (IgG).

A new technique is reported for the enhanced colon-metric detection of multiplexed hybridization onto porous membrane-based microarrays. This approach combines the use of horseradish peroxidase (HRP) as a label together with a chromogen substrate and a local production of the hydrogen peroxide required for substrate oxidation. This in situ production of coreagent is obtained using glucose oxidase (GOx) directly immobilized within the micro-array porous membrane mesh. The oxidation of glucose by the immobilized GOx produces hydrogen peroxide which itself enables the oxidation of TMB(3,3',5,5'-tetramethylbenzidine) by HRP and yields a blue precipitate on positive spots. Thanks to a coreagent overconcentration within the membrane, this design drastically surpasses the performances of the standard TMB/H2O2 kit used for peroxidase label detection. The obtained target limit of detection is then SO times lower (20 pM) than the one obtained with the standard kit approach, and the dynamic range expands at least one decade. Furthermore, the developed method was shown to compete well with the widely used alkaline phosphatase-BClP (5-bromo-4-chIoro-3-indolyl phosphate)/NBT (nitro blue tetrazolium chloride) readout while minimizing background signal. The method was finally successfully applied to the multiplex detection of single nucleotide polymorphisms (SNPs) in complex PCR samples. The background lowering was impacted here positively on the SNPs' detection by increasing the complementary noncomplementary signal ratio. [ABSTRACT FROM AUTHOR]

Peroxidase labeled streptavidin was immobilized onto the surface of bulk and clusterlike metal films at a distance controlled by a peptide chain with a length between 1.3 and 7.8 nm. Luminol chemiluminescence which occurred at peroxidase vicinity depends on the metal nanostructure. When peroxidase is attached on a bulklike film, chemiluminescence increases monotonously with the distance because of a decrease of the light emission quenching by metal. When peroxidase is attached on a clusterlike film, chemiluminescence undergoes a complex variation with the metal/catalyst distance evidencing a competition between the already mentioned quenching process and a nanostructure-induced catalysis enhancement. [ABSTRACT FROM AUTHOR]