Combinatorial Arrays Research Articles

Combinatorial Fluorescence XAFS Imaging of Manganese Complex Oxides

Abstract This report demonstrates that novel XAFS (X-ray absorption fine structure) microimaging, which does not require any spatial scans of the sample, can be used to evaluate the chemical states of combinatorial array libraries. The method has been applied to manganese–cobalt spinel MnCo2O4 prepared on the same substrate under different synthesis conditions. The valence numbers of Mn have been studied as a map by obtaining X-ray fluorescence images (1000 × 1000 pixels) as a function of the incident energy of synchrotron X-rays. The advantage of this method is that the whole substrate, i.e., all samples in the library, can be observed in a short time, typically ca. 10 min, which is equivalent to the measuring time for a single sample on the substrate when conventional scanning methods are employed.

From Proteins to Proteomics

During the second half of the 20th century, biochemistry and subsequently molecular biology blossomed into the core upon which all biological and biomedical sciences now depend. A major part of these closely related disciplines has been the study of the structure and function of proteins and the diverse biological functions that they perform. Early experimentation necessarily focused on individual entities, selected mainly for their activities, but as technology improved there developed a tendency to look at proteins as larger, interactive groups or clusters. Spurred by the recent exponential production of genomic sequence data for a rapidly increasing number of species, protein chemistry has now evolved into a new discipline, proteomics. In addition to embracing the methods and approaches that have served protein scientists well in the past, it includes, and is perhaps best defined by, high-throughput analyses based in large part on 2D gel electrophoresis, MALDI and ESI mass spectrometry and combinatorial arrays. Proteomic targets include the identification of all genome products and a mapping of their interactions and expression profiles. These hold great promise for the identification of disease markers and drug targets, but are not without their challenges and pitfalls.

High-Throughput Screening of Barrier and Adhesive Behavior of Polymeric Coatings

AbstractA combinatorial factory for the preparation and screening of polymeric coatings was developed. Coating formulations were prepared and coated using novel combinatorial techniques to obtain libraries of varying composition and thickness. The thickness of each film in a combinatorial array is rapidly determined via visible-light absorbance of optical dyes in conjunction with the Beer-Lambert relationship. These combinatorial libraries were then tested and screened using a variety of custom-made high-throughput methods. Combinatorial screening of oxygen and moisture vapor transmission rate, along with adhesive properties, are shown here. OTR and MVTR are determined using spectroscopic techniques. For adhesion, a spherical probe adhesive tester is able to generate parameters linked to tack, peel, and shear in one measurement. In addition to describing the testing methodology, benefits and shortcomings of these techniques will be highlighted.

Prediction of HPLC Conditions Using QSPR Techniques: an Effective Tool to Improve Combinatorial Library Design

The purification and characterization of compounds resulting from parallel synthesis or combinatorial chemistry has not yet been optimized to operate as a completely automated high-throughput process. Liquid chromatography/mass spectroscopy (LC/MS) is most commonly employed to carry out the characterization and identification of combinatorial compounds. This desired level of automation can only be accomplished if the separation conditions for every compound in the combinatorial array are known prior to the analysis. This study presents a quantitative structure retention relationship (QSRR) approach to predict the retention time of structurally diverse solutes under 75 different LC/MS conditions. Sixty-two compounds were analyzed using 15 commonly used HPLC columns under 5 different gradient conditions. The solute retention time was used as the dependent variable, and more than 1000 molecular descriptors were calculated for this compound set to generate QSRR models. After the elimination of highly correlated variables and those with zero variance, two different genetic algorithms were applied to identify the most significant descriptors. Following the variable selection, the identified descriptors were used to create QSRR models for each separation condition. The calculated stepwise multiple linear regression models have been proven to be statistically significant and highly predictive, with an average coefficient of determination (R2) of 0.86, an average cross-validated r2 of 0.62, r2 = 0.76, and an average F value of 27.29. The QSRR models can be used to design "analysis-friendly" library purification plates, in which compounds are arranged on the basis of their predicted separation condition and can also be used during the library design phase to flag compounds not amenable to the separation methods in use.

The Lovász Local Lemma and Its Applications to some Combinatorial Arrays

The Lovasz Local Lemma is a useful tool in the “probabilistic method” that has found many applications in combinatorics. In this paper, we discuss applications of the Lovasz Local Lemma to some combinatorial set systems and arrays, including perfect hash families, separating hash families, u-free systems, splitting systems, and generalized cover-free families. We obtain improved bounds for some of these set sytems. Also, we compare some of the bounds obtained from the local lemma to those using the basic probabilistic method as well as the well-known “expurgation” method. Finally, we briefly consider a “high probability” variation of the method, wherein a desired object is obtained with high probability in a suitable space.

Förster energy transfer in combinatorial arrays of selective doped organic light-emitting devices

Energy transfer in highly-efficient doped organic light-emitting devices (OLEDs) is described and discussed. The OLEDs include a hole transport layer (HTL) and an electron transport layer composed of an efficient blue emitter. A region of the HTL adjacent to the host blue-emitting layer was doped with an efficient guest red dye. The blue emitter-to-red dye energy transfer probability PHGη was determined by comparing the emission from the two fluorophores and its dependence on the applied field. PHGη decreases with increasing field, probably due to an increasing fraction of dye molecules which are positively charged, i.e., which trap a hole. It is also estimated that at fields as low as 0.4 MV/cm, ∼50% of the dye emission is due to trap emission rather than Förster energy transfer. The analysis yields a Förster energy transfer radius R0=33.5±3.5 Å.

A "catch and release" strategy for the parallel synthesis of 2,4,5-trisubstituted pyrimidines.

A resin capture and release strategy for making a combinatorial array of 2,4,5-trisubstituted pyrimidines is demonstrated by capturing beta-ketoesters and beta-ketoamides on a solid-supported piperazine. Through a cyclocondensation reaction, the solid-supported enaminone is reacted with several guanidines under heating or microwave irradiation affording the corresponding pyrimidines in good yield and chemical purity directly on solution. After this final step, the support can be effectively recycled.

Combinatorial chemistry methods for coating development V: generating a combinatorial array of uniform coatings samples

Combinatorial methods have proven to be valuable tools for the development of new compounds, catalysts, and electronic materials. We have developed a combinatorial factory capable of preparing and evaluating about 100 organic clear coatings in a day. Successful application of combinatorial methods to coatings development requires rapid and uniform preparation of a large number of diverse coatings. We have found that centrifugal spin casting is an effective method of producing an array of 48 coatings samples, each 9 mm diameter. The uniformity and flatness of each sample is dependent on specific process details such as protection from air currents and control of the evaporation process to avoid the “coffee ring effect”.

Diversified transcription initiation complexes expand promoter selectivity and tissue-specific gene expression.

Transcription initiation is a key regulatory step in the control of gene expression. Formation of a preinitiation complex at the right time and at the right promoter is a prerequisite to executing the correct programs of mRNA synthesis. This involves the interplay of many transcription factors and a combinatorial array of cis-regulatory DNA elements. Recent findings suggest that metazoan organisms have evolved specialized transcription initiation complexes and promoter-selectivity modules that direct coordinated regulation of functionally related gene networks. Here, we summarize corroborating evidence for a highly diversified core transcription machinery directing cell type-specific and gene-selective transcription. In multicellular organisms elaborate mechanisms have evolved to control the spatial and temporal patterns of transcription during growth, differentiation, and development. Transcriptional activation, one of the fundamental means of regulating gene expression in eukaryotes, is governed by an interconnected ensemble of multisubunit transcription factor complexes (Fig. 1; Lemon and Tjian 2000; Narlikar et al. 2002; Orphanides and Reinberg 2002). To assure the proper assembly of the transcription machinery, the components of the core transcriptional apparatus and the chromatin DNA template are subject to regulation. For instance, a variety of covalent histone and DNA modifications can influence whether a chromatin template is programmed to be transcriptionally active or silent (Strahl and Allis 2000; Berger 2002; Geiman and Robertson 2002). Also, numerous chromatin remodeling/nucleosome mobilizing activities catalyze the ATP-dependent deposition, removal, or sliding of nucleosomes to generate DNA templates that are accessible to the transcription machinery (Becker and Horz 2002). An enormous family of DNA sequence-specific transcriptional activators working in concert with various coregulatory factors drive the formation of active transcription initiation complexes (Lemon and Tjian 2000; Malik and Roeder 2000; Naar et al. 2001). It now seems clear that this highly regulated and coordinated assembly of active transcription complexes requires the interplay of a large number of transcription factors, multiple core promoter elements, and chromatin remodeling and modifying factors to properly position the preinitiation complex (PIC) at the start site of RNA synthesis (Burley and Roeder 1996; Roeder 1996; Lemon and Tjian 2000; Fig. 1). Until recently, the general transcription machinery that makes up the PIC was thought to be largely invariant when compared among different cell types within an organism. Although a large number of tissue-specific transcription factors have been described that control expression of cell type-specific genes, most of these have been enhancer-binding transcription factors. However, contrary to expectations, several studies have identified cell type-specific components of the core transcription machinery. Subsequently, an increasing number of cell type-specific and gene-selective homologs of basal transcription factors have been identified in metazoan organisms, including additional members of the TATA boxbinding protein (TBP) family such as TBP-related factors (TRFs) as well as numerous tissue-specific homologs of TBP-associated factors (TAFs). It now seems apparent that multicellular organisms have evolved cell type-specific components of the general transcriptional apparatus as a mechanism to accommodate more complex programs of tissue-specific and gene-selective transcription. Intriguingly, many of these functionally specialized core promoter factors are subunits of TFIID. This central component of the PIC harbors several transcription factors directly involved in recognizing and binding core promoter elements such as the TATA box, Initiator (Inr), and downstream promoter element (DPE). These findings suggest that metazoans have evolved functionally specialized transcription initiation complexes and promoter-selective modules, which might serve as eukaryotic factors (Gross et al. 1998; Losick 1998). In this review, we will summarize some recent advances in our understanding of how these specialized transcription initiation complexes might mediate proCorresponding author. EMAIL jmlim@uclink4.berkeley.edu; FAX (510) 643-9547. Article and publication are at http://www.genesdev.org/cgi/doi/10.1101/ gad.1099903.

Diastereoselective solution and multipin-based combinatorial array synthesis of a novel class of potent phosphinic metalloprotease inhibitors.

The solution-phase synthesis and resolution of new phosphinopeptidic building blocks containing a triple bond was realized in high yields and optical purities (units 3 a-d). The absolute configuration of the target compounds was unambiguously established by NMR studies. A post-assembly diversification strategy of these blocks was developed through 1,3-dipolar cycloaddition of a variety of in situ prepared nitrile oxides. This strategy led to the rapid and efficient diastereoselective preparation of a novel class of isoxazole-containing phosphinic peptides (peptides 5 a-i). Solid-phase version of this strategy was efficiently achieved on multipin solid technology, by developing a new protocol for the coupling of P-unprotected dipeptidic blocks with solid supported amino acids in a quantitative and diastereoselective manner. Optimization of dipolar cycloadditions onto pin-embodied phosphinic peptides allowed the convenient preparation of this new class of pseudopeptides. The crude phosphinic peptides (9 a-k) were obtained in high yields and purity as determined by RP-HPLC. Inhibition assays of some of these peptides revealed that they behave as very potent inhibitors of MMPs, outmatching previously reported phosphinic peptides, in terms of potency (K(i) in the range of few nM).

Combinatorial arrays and parallel screening for positive electrode discovery

Combinatorial techniques have been applied to the preparation and screening of positive electrode candidates for lithium batteries. This work describes the automated parallel synthesis of 64-electrode arrays using a Packard Multiprobe II liquid handling system. A cell was constructed with a single lithium reference–counter electrode and 64, three-millimeter-diameter working electrodes containing Li x Mn 2O 4 active material, PVdF–HFP binder and carbon black as a conducting additive. Eight duplicate electrodes, each of eight respective compositions, were deposited on the array and the mass fraction of carbon was varied in steps from 1 to 25%. The results showed a rapid increase in capacity at the percolation limit of 3% for most cells. Some groups of nominally identical cells showed random variations in capacity, especially at low carbon loadings. The overall result is a demonstration of advantages of the combinatorial concept, which were time-saving and an improved statistical significance of the results compared with on–off experiments.

Directional Characteristics of Resistivity Changes in Rock of Original Resistivity Anisotropy

AbstractAccording to pre‐designed patterns, multiple electrodes are arranged on the surfaces of four samples. Among them three samples have the original resistivity anisotropy which was not formed by ingredient of rock and one sample has that formed by ingredient of rock. The samples are fully saturated with water. The multiple electrodes are combined form arrays of resistivity changing anisotropy in different directions and spacing. The experiments of dynamic rock resistivity change are done with uniaxial compression, low confining triaxial compression and shear, and the variation of resistivity during the whole loading process is observed. The experimental results show that the samples of original resistivity anisotropy are essentially accordant with that of original resistivity isotropy in resistivity changing anisotropy. Namely, for the measuring‐points that are located at cracks or rupture bands, the results of resistivity changing anisotropy are good. The four directions deduced from four kinds of combinatorial equation sets of resistivity are essentially identical with each other and coincide with the orientations of cracks or main rupture bands approximately. For the measuring‐points without cracks or rupture bands passing through, either the four directions calculated by different combinatorial arrays of resistivity are inconsistent with each other, or the direction can not be determined, especially in the case that the crack plane is parallel to the measuring‐surface.

Differential receptors create patterns diagnostic for ATP and GTP.

Herein we report the combination of a library of resin-bound sensors along with a multicomponent sensor array. This novel combinatorial array sensor system shows selectivity for nucleotide phosphates in solution. The design of the anchored receptor includes a 1,3,5-trisubstituted-2,4,6-triethylbenzene scaffold coupled with peptide libraries. Each chemosensor is placed into a micromachined cavity within a silicon wafer, and the optical changes observed by a charged-coupled device result in near-real-time digital analysis of solutions. A colorimetric displacement assay was performed, and time-dependent imaging studies of the selected sensing ensembles result in a differential responses upon addition of adenosine 5'-triphosphate (ATP), adenosine 5'-monophosphate (AMP), or guanosine 5'-triphosphate (GTP). An advantage to this approach is that it creates an array of sensors that gives a fingerprint response for each analyte. Principal component analysis indicates that the library of chemosensors can differentiate between ATP, GTP, and AMP. On the basis of factor loading values, individual sensors from the library were sequenced to elucidate their chemical composition.

Combinatorial Fabrication and Studies of Small Molecular Organic Light-Emitting

ABSTRACTVarious combinatorial matrix arrays of UV-violet, white, and blue-to-red organic light-emitting devices (OLEDs), fabricated using a sliding shutter technique, are described. In the UV-violet devices, which contain a UV-violet emitting layer of 4,4′-bis(9-carbazolyl) biphenyl (CBP), the optimal radiance R and external quantum efficiency ηext were determined with respect to the thicknesses of the hole transporting layers. In the blue-to-red devices, which contained a blue-emitting layer of 4,4′-bis(2,2′-diphenyl-vinyl) -1,1′-biphenyl (DPVBi) and a red-emitting 5 wt.% dye-doped guest-host layer, the color of the devices evolved continuously from blue to red as the thickness of the doped layer increased from 0 to 35 Å. The (nominal) 2 Å-thick doped layer device exhibited the highest brightness L ∼ 120 Cd/m2 and external quantum efficiency ηext ∼4.4 % at a current density of 1 mA/cm2. In the white OLEDs, which were similar to the blue-to-red devices but with lightly doped emission layer, the highest brightness Lmax was over 74,000 Cd/m2; in all devices Lmax exceeded 50,000 Cd/m2. The maximum efficiencies were 11.0 Cd/A, 5.96 lm/W and 4.6% at 5.8 V, 0.6 mA/cm2, and 68 Cd/m2 in a 0.25 wt.%, 2 nm-thick doped layer device.

High-throughput purification of combinatorial arrays.

Performance and reproducibility of the Biotage Parallex, a high-throughput purification system, was evaluated using known standards. The results indicate that parallel purification is a robust technique for purifying large numbers of compounds. Results from one of the first libraries to be purified on the Biotage Parallex are presented and discussed. Since fractionation by UV can often result in a large number of fractions, threshold trigger versus yield and number of fractions was also investigated. This approach was used to purify an array of 4320 compounds, produced by an 11-step solid-phase synthesis in Irori MicroKans. Ninety-three percent of the compounds were successfully processed, with >90% having purity >95%.

Development of combinatorial chemistry methods for coatings: high-throughput optimization of curing parameters of coatings libraries.

An automated analytical system has been implemented for the high-throughput optimization of processing conditions such as curing parameters in fabrication of UV-cured automotive organic protective coatings. Selection of optimum process conditions of combinatorial arrays of coatings is essential to correlate the high-throughput screening and conventional processes and to achieve the desired physical properties of coatings. For monitoring of curing conditions of each coating in the array, a viscosity-sensitive fluorophore 4,4'-bis(2-benzoxazolyl)stilbene was incorporated into coating formulations. This fluorescence tagging approach permitted us to combine a gradient temperature heater and a UV curing system with the full capabilities of our high-throughput screening system, including generation of spectroscopic data and its analysis. This investigation demonstrated the possibility of rapid decoupling of temperature and radiation effects in curing of UV-curable coating formulations by using multiple coatings and process conditions at once. While the system described here was implemented for high-throughput optimization of temperature conditions of radiation curing of arrays of organic protective coatings for automotive applications, this system can be further applied for a variety of other applications where optimization of process parameters can be studied in situ or off-line using optical spectroscopic tools.

Microfluidic large-scale integration.

We developed high-density microfluidic chips that contain plumbing networks with thousands of micromechanical valves and hundreds of individually addressable chambers. These fluidic devices are analogous to electronic integrated circuits fabricated using large-scale integration. A key component of these networks is the fluidic multiplexor, which is a combinatorial array of binary valve patterns that exponentially increases the processing power of a network by allowing complex fluid manipulations with a minimal number of inputs. We used these integrated microfluidic networks to construct the microfluidic analog of a comparator array and a microfluidic memory storage device whose behavior resembles random-access memory.

Structure-based combinatorial library design: discovery of non-peptidic inhibitors of caspases 3 and 8.

Structure-based design of a combinatorial array was carried out in order to identify non-peptidic thiomethylketone inhibitors of caspases 3 and 8. Five compounds from the designed array were active against caspase 3, and two were active against caspase 8. A 2.5-A resolution co-crystal structure of caspase 3 and a thiomethylketone array member is reported. The structure-based design strategy has proved useful for identifying caspase inhibitors.

Ultrafast algorithm for designing focused combinational arrays

A novel greedy algorithm for the design of focused combinatorial arrays is presented. The method is applicable when the objective function is decomposable to individual molecular contributions and makes use of a heuristic that allows the independent evaluation and ranking of candidate reagents in each variation site in the combinatorial library. The algorithm is extremely fast and convergent and produces solutions that are comparable to and often better than those derived from the substantially more elaborate and computationally intensive stochastic sampling techniques. Typical examples of design objectives that are amendable to this approach include maximum similarity to a known lead (or set of leads), maximum predicted activity according to some structure-activity or receptor binding model, containment within certain molecular property bounds, and many others.

Combinatorial screening of enzyme activity by using multiplexed capillary electrophoresis.

Efficient and comprehensive screening of enzyme activity was accomplished in a combinatorial array of 96 reaction microvials. Quantitation of the extent of the reaction at well-defined time intervals was achieved by using 96-capillary array electrophoresis coupled with a multiplexed absorption detector. Capillary electrophoresis provides high separation resolution to isolate the product from the reactants. Absorption detection provides universal applicability to combinatorial screening. For the conversion of NADH to NAD+, the catalytic activity of LDH was confirmed to be the highest at pH 7. This scheme should be useful for high-throughput drug discovery, clinical diagnosis, substrate binding, as well as combinatorial synthesis.