Toward a microchip-based solid-phase extraction method for isolation of nucleic acids

A silica-based solid-phase extraction system suitable for incorporation into a microchip platform (nu-total analytical system; nu-TAS) would find utility in a variety of genetic analysis protocols, including DNA sequencing. The extraction procedure utilized is based on adsorption of the DNA onto bare silica. The procedure involves three steps: (i) DNA adsorption in the presence of a chaotropic salt, (ii) removal of contaminants with an alcohol/water solution, and (iii) elution of the adsorbed DNA in a small volume of buffer suitable for polymerase chain reaction (PCR) amplification. Multiple approaches for incorporation of this protocol into a microchip were examined with regard to extraction efficiency, reproducibility, stability, and the potential to provide PCR-amplifiable DNA. These included packing microchannels with silica beads only, generating a continuous silica network via sol-gel chemistry, and combinations of these. The optimal approach was found to involve immobilizing silica beads packed into the channel using a sol-gel network. This method allowed for successful extraction and elution of nanogram quantities of DNA in less than 25 min, with the DNA obtained in the elution buffer fraction. Evaluation of the eluted DNA indicated that it was of suitable quality for subsequent amplification by PCR.     

A microchip-based proteolytic digestion system driven by electroosmotic pumping

Microchip-based proteomic analysis requires proteolytic digestion of proteins in microdevices. Enzyme reactors in microdevices, fabricated in glass, silicon, and PDMS substrates, have recently been demonstrated for model protein digestions. The common approach used for these enzyme reactors is employment of a syringe pump(s) to generate hydrodynamic flow, driving the proteins through the reactors. Here we present a novel approach, using electroosmotic flow (EOF) to electrokinetically pump proteins through a proteolytic system. The existence of EOF in the proteolytic system packed with immobilized trypsin gel beads was proven by imaging the movement of a neutral fluorescent marker. Digestions of proteins were subsequently carried out for 12 min, and the tryptic peptides were analyzed independently using capillary electrophoresis (CE) and MALDI-TOF mass spectrometry (MS). The results from CE analysis of the tryptic peptides from the EOF-driven proteolytic system and a conventional water bath digestion were comparable. MALDI-TOF MS was used to identify the parent protein and the tryptic peptides using MS-Fit database searching. The potential utility of the EOF-driven proteolytic system was demonstrated by direct electro-elution of proteins from an acrylamide gel into the proteolytic system, with elution and tryptic digestion achieved in a single step. The EOF-driven proteolytic system, thus, provides a simple way to integrate protein digestion into an electrophoretic micro total analysis system for protein analysis and characterization.     

On three problems for sequences

If ξ∈ (0,1) and A=a_n, n?? is a sequence of real numbers define S_n(ξ,A)∶=Σ{a_k∶:k=[nξ]+1 to n}, n??, where [x] is the greatest integer less than or equal to x. In the theory of regularly varying sequences the problem arose to conclude from the convergence of the sequence S_n (ξ,A), n??, for all ξ in an appropriate set K of real numbers, that the sequence a_n, n??, converges to zero. It was shown that such a conclusion is possible if K={ξ,1?ξ} with ξ∈ (0,1) irrational. Then the following three questions were posed and will be answered in this paper:

1. does the convergence of S_n (ξ,A), n??, for a single irrational number ξ imply a_n→0.
2. does the convergence of S_n(ξ,A), n??, for finitely many rational numbers ξ∈ (0, 1) imply a_n→0.
3. does the convergence of S_n (ξ,A), n??, for all rational numbers ξ∈ (0,1) imply a_n→0?

     

On the Rayleigh-Plateau instability

In this paper,we study the surface instability of a cylindrical pore in the absence of stress. This instability is called the Rayleigh-Plateau instabilty. We consider the model developed by Spencer et ...     

DNA purification using dynamic solid-phase extraction on a rotationally-driven polyethylene-terephthalate microdevice

We report the development of a disposable polyester toner centrifugal device for semi-automated, dynamic solid phase DNA extraction (dSPE) from whole blood samples. The integration of a novel adhesive and hydrophobic valving with a simple and low cost microfabrication method allowed for sequential addition of reagents without the need for external equipment for fluid flow control. The spin-dSPE method yielded an average extraction efficiency of ∼45% from 0.6 μL of whole blood. The device performed single sample extractions or accommodate up to four samples for simultaneous DNA extraction, with PCR-readiness DNA confirmed by effective amplification of a β-globin gene. The purity of the DNA was challenged by a multiplex amplification with 16 targeted amplification sites. Successful multiplexed amplification could routinely be obtained using the purified DNA collected post an on-chip extraction, with the results comparable to those obtained with commercial DNA extraction methods. This proof-of-principle work represents a significant step towards a fully-automated low cost DNA extraction device.     

Synthesis,Antibacterial, and Antioxidant Evaluation of Novel Series of Condensed Thiazoloquinazoline with Pyrido,Pyrano, and Benzol Moieties as Five- and Six-Membered Heterocycle Derivatives

A novel synthesis of thiazolo[2,3-b]quinazolines 4(a–e), pyrido[2′,3′:4,5]thiazolo[2,3-b]quinazolines {5(a–e), 6(a–e), and 7(a–e)}, pyrano[2′,3′:4,5]thiazolo[2,3-b]quinazolines 8(a–e), and benzo[4,5]thiazolo[2,3-b]quinazoloine9(a–e) derivatives starting from 2-(Bis-methylsulfanyl-methylene)-5,5-dimethyl-cyclohexane-1,3-dione 2 as efficient α,α dioxoketen dithioacetal is reported and the synthetic approaches of these types of compounds will provide an innovative molecular framework to the designing of new active heterocyclic compounds. In our study, we also present optimization of the synthetic method along with a biological evaluation of these newly synthesized compounds as antioxidants and antibacterial agents against the bacterial strains, like S. aureus, E. coli, and P. aeruginosa. Among all the evaluated compounds, it was found that some showed significant antioxidant activity at 10 μg/mL while the others exhibited better antibacterial activity at 100 μg/mL. The results of this study showed that compound 6(c) possessed remarkable antibacterial activity, whereas compound 9(c) exhibited the highest efficacy as an antioxidant. The structures of the new synthetic compounds were elucidated by elemental analysis, IR, ¹H-NMR, and ¹³C-NMR.     

The Roles of Composition and Mesostructure of Cobalt-Based Spinel Catalysts in Oxygen Evolution Reactions

By using the crystalline precursor decomposition approach and direct co-precipitation the composition and mesostructure of cobalt-based spinels can be controlled. A systematic substitution of cobalt with redox-active iron and redox-inactive magnesium and aluminum in a cobalt spinel with anisotropic particle morphology with a preferred 111 surface termination is presented, resulting in a substitution series including Co₃O₄, MgCo₂O₄, Co₂FeO₄, Co₂AlO₄ and CoFe₂O₄. The role of redox pairs in the spinels is investigated in chemical water oxidation by using ceric ammonium nitrate (CAN test), electrochemical oxygen evolution reaction (OER) and H₂O₂ decomposition. Studying the effect of dominant surface termination, isotropic Co₃O₄ and CoFe₂O₄ catalysts with more or less spherical particles are compared to their anisotropic analogues. For CAN-test and OER, Co³⁺ plays the major role for high activity. In H₂O₂ decomposition, Co²⁺ reveals itself to be of major importance. Redox active cations in the structure enhance the catalytic activity in all reactions. A benefit of a predominant 111 surface termination depends on the cobalt oxidation state in the as-prepared catalysts and the investigated reaction.     

Fully differential cross sections for photo-double-ionization of D2

We report the first kinematically complete study of the four-body fragmentation of the D2 molecule following absorption of a single photon. For equal energy sharing of the two electrons and a photon energy of 75.5 eV, we observed the relaxation of one of the selection rules valid for He photo-double-ionization and a strong dependence of the electron angular distribution on the orientation of the molecular axis. This effect is reproduced by a model in which a pair of photoionization amplitudes is introduced for the light polarization parallel and perpendicular to the molecular axis.     

Direct capillary electrophoretic detection of carbohydrate-deficient transferrin in neat serum

Transferrin, an iron transport protein found in serum and cerebrospinal fluid, is known to be microheterogeneous with respect to its carbohydrate and sialic acid content. The forms of transferrin deficient in sialic acid and/or carbohydrate, termed carbohydrate-deficient transferrin (CDT), have been of clinical interest for almost two decades as a result of the initial finding that elevated CDT concentrations are associated with chronic, excessive alcohol abuse. We demonstrate the utility of capillary electrophoresis for examining the CDT sialoform profile via the direct electrophoresis of serum. The need for negligible preelectrophoresis sample preparation and absence of postelectrophoresis processing dramatically decreases analysis time compared to slab gel-based separations. Using a fluorocarbon-coated capillary containing a hydroxyethyl cellulose/borate buffer, the high resolution separation of serum components is effected in less than 30 min. Under these conditions, the beta region proteins (including transferrin) are well resolved from the alpha-2 and gamma zone proteins in a window where the individual transferrin sialoforms can be detected. The usefulness of this method is demonstrated with the electrophoresis of serum from subjects known to be either non-alcoholic and alcoholic.