Fabrication of a totally renewable off-channel amperometric platform for microchip electrophoresis.

In this approach, a novel method to fabricate an integrated amperometric platform used in off-channel electrophoresis has been introduced. A simple screen printed protocol combining a wet etching procedure was used to define the pattern on a glass substrate, and whole electrodes were constructed by filling the conductive carbon ink into the etched cavities. A simple Teflon tape was used to align this platform with the micro-channel, and the variation of reassembling of this device can be down to 2.2% without the assistance of microscope. This device was characterized by dopamine (DA) and catechol (CA), and the width of half peak is around 4s, even a 100 µm double T shape injection design and a 550 µm working electrode were used in this work. Under the optimum condition, this device possesses a low background with a noise level of 1.4 pA (peak to peak). The linear range for DA and CA are 0.1-100 µM (R = 0.998) and 0.2-200 µM (R = 0.996) with a theoretical plate number of 1.57 × 10(4) and 3.46 × 10(4) (plate/m), respectively.

Low-potential amperometric determination of purine derivatives through surface oxide regeneration method.

This article described a novel amperometry which can be used for determination of purine derivatives including uric acid, xanthine, hypoxanthine, guanine, and adenine without surface contamination. By applying a constant potential of -0.125 V (vs. Ag/AgCl) in a flow injection system, the chelating capability of these purine derivatives converts the cuprous oxide layer into a soluble complex. This behavior would dissolve the passive oxide layer and expose the bottom copper layer to the solution, subsequently; an oxidation current which attributed to the regeneration of the original cuprous oxide layer is used to reflect the concentration of these purine derivatives. In a 50mM phosphate buffer, pH 7.0, this approach provides a high sensitivity with LOQ of sub-micro molar level of five purines and high stability with a RSD of 2.5% for 10 µM xanthine (N=12). This method does not suffer from most biological species including ascorbic acid, acetaminophen, creatine, dopamine, sarcosine, ammonium ion, chloride ion, and urea at equal or higher than its physiological concentration.

Amperometric determination of NADH with Co3O4 nanosheet modified electrode.

In this work, we have developed a simple and reliable cobalt oxide (Co3O4) based amperometric sensor for the determination of NADH. A sheet shape Co3O4 nanooxide was synthesized by the CTAB assisted hydrothermal technique and was characterized by SEM and XPS. Owing to the redox property of Co3O4, the operating potential of NADH can be significantly reduced from 0.7 down to 0.1 V. Compared to a commercial Co3O4 nanoparticle modified electrode, this nanosheet form cobalt oxide possesses a rapid background subsiding characteristic and a low residual current. This scheme was conducted on a flow injection system with a constant operating potential of 0.1 V (vs. Ag/AgCl, 3 M) in a 0.2 M phosphate buffer at pH 6.0. A suitable linear range from 10 to 100 µM (R=0.999) with a detection limit of 4.25 µM (S/N=3) was obtained. The RSD for 20 successive measurements of 75 µM NADH is only 1.4%, which indicates a high stability and no contamination during NADH oxidation. This scheme did not suffer from conventional antioxidants, including dopamine, uric acid, epinephrine, serotonin, histamine, and 4-acetaminophen, except ascorbic acid. Thus, an ascorbate oxidase was introduced to remove the ascorbic acid before the sample was injected into the flow injection analysis system. After this simple pretreatment, the influence of ascorbic acid was eliminated, successfully.

A novel structural specific creatinine sensing scheme for the determination of the urine creatinine.

In this work, a highly structural dependent amperometric scheme was proposed for the determination of creatinine without enzymatic assistance. The principle of this novel method is based upon the formation of a soluble copper-creatinine complex on the copper electrode surface. Subsequently, an oxidative current from the regeneration of the surface oxide layer is monitored and it is proportional to the concentration of the creatinine. This scheme can be conducted at potential of -0.1 V (vs. Ag/AgCl, 3 M) in phosphate buffer (pH 7). A typical calibration plot from 25 µg/dL to 1.5 mg/dL (R(2)=0.997) with a detection limit of 6.8 µg/dL (S/N=3) is achieved. The relative standard deviation of 21 successive injections of 0.2 mg/dL creatinine is 0.018. Under the optimal conditions, the frequently encountered biological interferences at physiological or higher concentration were investigated. Only uric acid revealed an obvious interference (298.1%). However, a Nafion(®) coated copper plating electrode shows a successful decrement of the interference of the uric acid with slightly decreased sensitivity of creatinine. The feasibility of this scheme for further clinical application is demonstrated by both HPLC and FIA to evaluate the creatinine concentration in a urine sample.

Affinity chromatography of DNA on nonporous copolymerized particles of styrene and glycidyl methacrylate with immobilized polynucleotide.

Nonporous particles of microsize were prepared by the dispersion polymerization of styrene and glycidyl methacrylate and chemically modified to introduce amino groups on the surface by grafting with either hexamethylenediamine or N-methyl-1,3-propanediamine. Aminated particles were then coupled with phosphorylated single-stranded polynucleotides at the 5'-end through covalent linkages. The affinity columns packed with these prepared polynucleotide-immobilized particles effectively retained single-stranded DNA, which could base-pair with the immobilized sequence. Bound DNAs could be eluted to yield a sharp peak by using an aqueous solution of 0.4M NaOH. The nonspecific adsorption due to the electrostatic interaction between the polynucleotide and the residual amino groups on the particle surface via the amination with hexamethylenediamine was significant and could only be reduced by using a high salt (NaCl) concentration. A higher salt concentration in the elution solution could result in a portion of complementary polynucleotide eluted in the nonretained fraction. However, the nonspecific adsorption of polynucleotides was insignificant in the column packed with DNA-immobilized particles prepared via amination using N-methyl-1,3-propanediamine. The column was effective for microanalysis of sequence-specific DNA.

Influence of Surface Hydrophobic Groups on the Adsorption of Proteins onto Nonporous Polymeric Particles with Immobilized Metal Ions.

Iminodiacetic acid (IDA) and octyl moieties were covalently bound on nonporous particles, which were prepared from dispersion polymerization of methyl methacrylate and glycidyl methacrylate. After being charged with copper ions, the IDA-bound particles could specifically adsorb deoxyribonuclease I (DNase I) through the affinity interaction between protein and immobilized metal ion. A mixed-ligand (metal-chelate and octyl-bound) support was obtained after hydrophobic (octyl) groups were also introduced to the particle surface. The affinity adsorption of DNase I on the copper-IDA chelate was influenced by interaction between the protein and the bound octyl group. Both the affinity and the hydrophobic interactions could be well described by the Langmuir isotherms. The equilibrium adsorption constants were estimated separately to be 0.96 and 0.50 liter g(-1) for affinity and hydrophobic bindings, respectively. For binding on mixed-ligand support, the adsorption constant was 0.45 liter g(-1). It was evident that both affinity and hydrophobic interactions are involved in the adsorption of proteins onto mixed-ligand particles. Desorption of the inactive proteins from the support was possible by increasing the hydrophobicity of the solution. Copyright 2001 Academic Press.