This paper describes some basic test schemes of piezoelectric quartz crystal microbalance (QCM)-based immunoassay. These include one-step direct assay, one-step indirect assay, two-step sandwiched assay, displacement assay, and mass-amplified assay. Applications of these designs in the fields of clinic diagnosis, food analysis, environmental analysis, and veterinary diagnosis are reviewed. In addition, some critical issues of QCM immunoassay are also dealt with, including the evaluation of liquid phase and gas phase detection, and the biomolecules’ coating techniques, non-specific adsorption, and regeneration of used crystals.

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The hydrogen-bond formation of tris(pentane-2,4-dionato)cobalt(III) (Co(acac)₃) and tris(8-quinolinolato)cobalt(III) (Co(q)₃) with a series of chlorinated phenols has been studied by IR in chloroform and carbon tetrachloride at different temperatures ranging from 288 to 308 K. The IR spectra show clear evidence of hydrogen bonding between the chelates and the chlorinated phenols. From an equilibrium analysis, hydrogen-bonded complexes were found to be MB₃·nHA, where MB₃ and HA stand for the metal chelate and phenol, respectively, and n=1 - 3. The overall formation constants, βn (βn=[MB₃·nHA]/[MB₃][HA]ⁿ ), were determined and considered in relation to acid-dissociation constants of the phenols. From the temperature dependence of the formation constants, the thermodynamic parameters were determined. Additionally, the validity of the obtained enthalpy was ascertained by calorimetric titration in the Co(q)₃-3,5-dichlorophenol system. The negative ΔH⁰ and ΔS⁰ values obtained suggest the formation of a very strong type of hydrogen-bonded complexes, and that the thermodynamics of hydrogen-bond formation is dominated by ΔH⁰. Considering the formation constants and the thermodynamic parameters, Co(q)₃ has a hydrogen-bonding ability stronger than that of Co(acac)₃.

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A new immunoassay using chemiluminescence detection of dyestuff-containing liposomes as a labeling reagent is proposed. Human serum albumin labeled with Eosin Y containing-liposomes was employed as a labeling reagent for an immunoassay. After the immune reaction was carried out in the presence of an antibody-immobilized glass bead, the reactant solution was subjected to the capillary electrophoresis-hemiluminescence detection system. That is, the bound/free separation was conducted by use of glass beads, also, other coexisting compounds which might influence chemiluminescence detection were easily separated from the labeled human serum albumin by capillary electrophoresis. The labeled human serum albumin in the reactant solution was detected with high sensitivity. The amount of the labeled human serum albumin showed a good relationship to that of human serum albumin as an analyte through immune reaction. Human serum albumin could be determined over the range of 1×10^-6 - 5×10^-4 M. The present method was also applicable to the determination of protein in a serum sample without being interfered with by coexisting constituents.

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Simultaneous determinations of the concentration and molecular weight of humic substances in concentrated and unconcentrated environmental water samples by gel chromatography with a fluorescence detector were carried out. The calibration curves of humic acid and fulvic acid were linear in the concentration range of 0.1 to 10 mg/l. The relative standard deviations of humic substances were less than 10%. The concentrations of aquatic humic substances in rivers were high in the summer and low in the winter. The molecular weight of most humic substances in Katsura, Uji, Kidzu and Yodo Rivers was estimated to be about 3000 - 10000. The ratio of the higher molecular weight of humic substances was larger in the warmer season than that in the cooler season. The main origin of aquatic humic substances in river water may be humic substances from soils around the rivers.

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The dependence of the enantioselectivity and enantiomer migration order (EMO) on the chirality and stereo-conformation of ligands used for the chiral selectors of Cu(II) complexes was investigated. It was interestingly observed that EMOs were reversed not only by a change of the ligand chirality of selector, but also by the trans- or cis-conformation of a 4-hydroxy substituent in 4-hydroxyproline. In the presence of the sodium dodecyl sulfate (SDS) micellar phase, the EMOs were reversed when both the bidentate and tridentate ligands of the proline family were used. The resolution (Rs) of enantiomers decreases when the cis-hydroxy substituent exists in D- or L-hydroxyproline. SDS micellar phase improves the resolution of amino acid enantiomers. The relationships among the EMO, ligand chirality, complex stability and stereo-selectivity are discussed.

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We conducted a study for the separation of four catechins ((-)-epicatechin (EC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECG) and (-)-epigallocatechin gallate (EGCG)) in tea by HPLC with an electrochemical detector at different temperatures (25 - 50°C). Various solvent compositions of mixed solvents of water, acetonitrile (AN) and ethyl acetate (EA) were used as eluting solvents. The compositions of the solvents were as follows: water, 90 - 70% (v/v); AN, 10 - 30% (v/v); EA, 0 - 5% (v/v). The binary mixtures of water (90 - 70% (v/v)) and AN (10 - 30% (v/v)) were not able to separate the peaks of EC and EGCG under all of the different solvent compositions. However, ternary mixed-solvents containing a small amount of EA (3 - 5% (v/v)) with water (85 - 83% (v/v)) and AN (12% (v/v)) separated four catechins sufficiently, especially the peaks of EC and EGCG, within a 10 min elution time at 40°C. The effect of the solvent composition is discussed in terms of the E_T^N and D_II,I values, solvent structure and extraction constants of catechins into EA. The formation of a pseudo stationary phase in which ODS is partly covered by EA is proposed for the enhanced separation of EC and EGCG in the presence of EA. Catechins in three kinds of tea were determined under an optimum condition.

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The reaction of four aromatic aldehydes including p-dimethylaminobenzaldehyde (p-DMABA) with serum protein has been studied. The aromatic aldehydes gradually reacted with serum globulin with the irradiation of a fluorescent lamp at room temperature to form a colored product with an absorption maximum at about 520 to 635 nm with high molar absorptivity, while only slightly reacting with serum albumin. The reaction rate increased as the intensity of light increased, and at about 3000 luxes the reaction completed within 45 min after the reaction started. Of the four aromatic aldehydes evaluated, the colored product of p-DMABA indicated the most excellent spectral characteristics. Based on these results, this aromatic aldehyde was applied to the detemination of the serum total globulin concentration. The measurement values by the proposed method, using a γ-globulin solution as a standard material, correlated well with those by the method of Goldenberg and Drewes using glyoxylic acid (r = 0.983), the combined biuret-BCG method (r = 0.949) and moderately with the combined biuret-cellulose acetate electrophoresis method (r = 0.899).

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A very simple, selective and sensitive method is developed for the spectrophotometric determination of Cr^VI and/or V^V based on their reactions with perphenazine to instantaneously give a red colored product exhibiting a maximum absorbance at 526 nm. Following the recommended procedure, chromium and vanadium can be determined with linear calibration graphs up to 0.40 and 1.00 µg ml^-1 and detection limits of 3 and 5 ng ml^-1, respectively. The molar absorptivities are 1.87×10⁴ and 1.20×10⁴ l mol^-1 cm^-1 with Sandell sensitivity indexes of 2.8 and 4.2 ng cm^-2 for Cr and V, respectively. Analyses of mixtures of Cr^VI and V^V in the ratios of 1:100 to 100:1, following the recommended procedure gave recoveries of 97 - 102% with relative standard deviations of ≤1.4% for both Cr^VI and V^V. The total Cr and V was determined first; then, V was determined after addition of As^III to reduce Cr^VI that was calculated by the difference. Statistical treatment of the analytical results did not detect any systematic error and showed the high accuracy and precision of the method. The unique selectivity and sensitivity of the method allowed its direct application to the determination of Cr and V in complex matrices of certified reference materials and synthetic mixtures. The results obtained are in excellent agreements with the nominal values.

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A chelate of beryllium acetylacetonate, Be(acac)₂, was formed from beryllium(II) (1.0 - 40.0 ng) in drinking water (100 ml) containing acetylacetone (0.2%(v/v), 7.0 - 14.0 ml) in ammonium acetate buffer (14 - 28 mmol) together with a slightly acidic solution (0.2%(v/v) of HCl, 3.0 ml) at pH 5.5. The Be(acac)₂ chelate was preconcentrated on a Sep-Pak C₁₈ cartridge and was eluted with methanol (2.00 ml). A portion (20 µl) was introduced into a graphite cuvette and was atomized according to a temperature program. The method detection limit (3σ) for beryllium(II) was 2.3 ng/l; the calibration graph was linear up to 400 ng/l. Accuracies of 97.0 - 101% were obtained when testing a quality control (QC) water sample (HPS #290608) and a synthetic natural water standard (Standard Reference Material 1643d) with a relative standard deviation (n=4) within 2.3%. The proposed method could be applied to the measurement of beryllium vapor in air with a detection limit of 0.23 ng.

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Determination of As by direct electrothermal atomization atomic absorption spectrometry with a pyropytically-coated graphite furnace treated with W by a single drop treatment technique was reported. The treated PG furnace made it possible to introduce a 100 µl aliquot of sample solution into the furnace with a high precision (RSD <4%). Use of a Pd-modifier made it possible to enhance the precision (RSD <3%) and the sensitivity. A good linearity (correlation factor >0.99) between integrated absorbance and injection volume was observed in the range 0 - 100 µl. The calculated detection limit and characteristic mass (sensitivity) for integrated absorbance were 22 ng l^-1 and 16 pg and for maximum absorbance were 45 ng l^-1 and 13 pg for a 100 µl injection with the Pd modifier at the optimum ashing temperature of 800°C. An interference study for environmental matrices was also carried out. The recoveries of 2 and/or 4 µg l^-1 of As added to various water samples, such as snow, cloud, river and tap waters, were 97 - 104%.

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The large synergistic effect of 3,5-dichlorophenol (Hdcp) in the extraction of gallium(III) with 2-methyl-8-quinolinol derivatives (HA), such as 2-methyl-8-quinolinol (HMQ), 2-methyl-5-ethoxymethyl-8-quinolinol (HMO₂Q), 2-methyl-5-butyloxymethyl-8-quinolinol (HMO₄Q), and 2-methyl-5-hexyloxymethyl-8-quinolinol (HMO₆Q), from a weakly acidic solution into heptane and chloroform was investigated. The acid-dissociation constants of HA in an aqueous solution, the distribution constants of HA between heptane and water, and the association constants between HA and Hdcp in heptane were determined. The extractability of gallium(III) with HA increased with an increase in the hydrophobicity of HA. The extractability of gallium(III) with HA was remarkably enhanced upon the addition of Hdcp. The synergistic effect of Hdcp was quantitatively evaluated in terms of both a mixed ligand complex formation of Ga(dcp)(A)₂ in an aqueous phase and the formation of an association-complex in an organic phase. The mixed-ligand complex formation and the association constant were determined at I = 0.1 M (H,Na)ClO₄ (1 M = 1 mol dm^-3) and 25°C. The compositions of gallium(III)-HA complexes extracted in both the absence and presence of Hdcp were assigned to be Ga(OH)(A)₂ and Ga(dcp)(A)₂·Hdcp, respectively. Moreover, the association constants of the synthesized Ga(OH)(MQ)₂ with Hdcp in anhydrous chloroform were also determined by FT-IR spectrometry and the association-complexs were to be Ga(OH)(MQ)₂·mHdcp (m = 1, 2).

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An overview is given on the recent development of combined flow injection (FI) capillary electrophoresis (CE) systems. The basic principles are treated and the advantages of the combined system, including improved throughput and reproducibility, and the potentials of incorporating various on-line separation and preconcentration systems, including filtration, dialysis, gas diffusion, and column sorption, are discussed and illustrated by applications.

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The structure characterization of proteins or enzymes by STM on electrochemically prepared HOPG surface studied in this laboratory is reviewed. The serial structures of Hb were observed. The differences between the denaturation and inactivation of HRP were investigated by in situ and ex situ STM. The structural variation of Hb in an organic solvent was imaged while protein denaturation was easily observed in a polar solvent.

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A molecular imprinted polymer was prepared using trimethoprim as the template molecule and methacrylic acid as the functional monomer by a non-covalent method. The powders of the polymer were coated on one electrode of a bulk acoustic wave (BAW) device to construct a biomimetic sensor for the determination of trimethoprim. In absolute ethanol, the sensor exhibited good selectivity and sensitivity. In the range of 6.0×10^-8 - 6.7×10^-5 M, there was a linear relationship between log(-Δf) and log C. The detection limit was 4.0×10 -8 M. When treated by heat, heavy metal salt, acid, etc., the sensor still exhibited long-time stability and resistance to such harsh chemical environments. Satisfactory results of a real sample assay were obtained by the proposed method.

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In this study, a second-order standard addition method based on alternating trilinear decomposition (ATLD-SOSAM) was developed. It was applied to second-order HPLC-DAD data and compared to methods employing direct trilinear decomposition (DTLD) and PARAFAC. The results show that ATLD-SOSAM is slightly superior to both DTLD-SOSAM and PARAFAC-SOSAM.

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A sample pretreatment by microwave-assisted extraction (MAE), microwave-assisted alkaline degradation (MAAD) and microwave-assisted saponification prior to the determination of PAHs, PCBs, triazines and cholesterol in diverse samples using GC, GC-MS and HPLC has been developed. When MAAD was coupled with MAE, the degradation of chlorinated pesticides and the extraction of PCBs analytes in the samples could be carried out simultaneously. Likewise, when MAAD and MAS were coupled with MAE, the degradation, saponification and extraction could be carried out simultaneously. In addition, a microwave-assisted derivatization (MAD) of fatty acids in edible oil, plasma and shark cartilage has been developed. The analytical results are satisfactory. The microwave-assisted sample pretreatment methods are rapid, labor and solvent saving, moreover, non-chlorinated organic solvents can be used.

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Thiocyanide ion adsorption on Pt and Ni electrode surfaces was investigated by confocal Raman microscopy. The effects on the adsorption behavior of SCN^- by the electrolyte anion, concentration and applied potential were studied. The potential has a significant effect on the frequency shift of SCN^- as a result of the electrochemical Stark effect. The adsorbability of the electrolyte anion has a significant influence on the surface Raman intensity of SCN^-; the intensity follows the order Cl^-<ClO₄^-<F^-. It can be explained in terms of competitive adsorption between the SCN^- and the anion. It has been shown that surface Raman spectroscopy can be a sensitive tool for the in-situ analysis of the surface species and its configuration, such as surface complexes and films on transition metal electrodes.

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A chemically modified electrode was successfully prepared by self-assembling bi-(2-aminoethyl)-aminodithiocarboxyl acid (BADTC) on a gold electrode and its further derivatization to form a monolayer of nickel hexacyanoferrate (NiHCF). Its electrochemical characteristics were investigated by cyclic voltammetry and an AC impedance spectroscopy technique. The electrochemical behavior of this modified electrode greatly depended on the kinds and concentrations of cations. Among the studied cations, including Li⁺ , K⁺ , Na⁺ , NH₄⁺ , Ca²⁺ and Mg²⁺ etc., K⁺ was found to be the most appropriate ion to participate in the electrochemical process of this BADTC-NiHCF film modified electrode, as a counterion. The heterogeneous rate constants of this film modified electrode were 0.95, 4.97 s^-1 in 0.001 and 0.01 mol L^-1 KCl solutions, respectively, obtained from the AC impedance. Moreover, this monolayer film modified electrode showed a good molecular-recognition ability for K⁺ and NH₄⁺.

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The matrix effects on analyte vaporization and ICP excitation conditions in direct sample insertion-inductively coupled plasma atomic emission spectrometry were studied. Zn was used as a test element and NaCl as the matrix. A slow insertion speed of the sample probe was used to allow partial separation in time of the vaporization of the volatile Zn species and the NaCl matrix. Temporal studies of the Zn emission intensity indicate that ZnCl₂ starts to vaporize during sample probe insertion in the presence of the NaCl matrix. The Zn intensity is reduced because the plume of Zn vapor is not concentrated in the central channel of the ICP. Temporal profiles of the intensity ratio of ZnII/ZnI also shows that introduction of a large quantity of NaCl into the ICP has little effect on the ICP excitation conditions. The ICP is, however, significantly cooled due to heat loss to the sample probe.

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Dissolved species of lanthanum in the solutions which contained ethylenediamine tetraacetic acid (EDTA) and L-tryptophan, respectively, were evaluated by electrospray ionization mass spectrometry. The stability of the species of La complexes during the cation-exchange chromatographic separation process was discussed. The results indicated that the speciation of lanthanum in the solution was remarkably influenced by the dissociation of the La species, which is in turn depended on the cation-exchange column selected.

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A rapid and simple homogeneous fluorescence PCR assay was developed for the clinical diagnosis of infectious diseases based on a molecular beacon. The established method could reproducibly detect Mycobacterium tuberculosis at the 10 bacteria/mL level. The analytical specificity was tested with 14 strains of mycobacterium, four unrelated bacteria and 220 negative samples; no false positive results were obtained. A blind test was also performed to evaluate its performance in Mycobacterium tuberculosis diagnosis. The results showed that both the clinical sensitivity and the specificity were 100%, and that the detection limit was in the range of 1 - 10 bacteria/ml. A clinical study with 466 patient samples demonstrated that fluorescence PCR assay correlated well with smear (93.6%) and culture (98.4%) methods for positive samples. However, fluorescence PCR could detect positive samples (62.9%) more than smear (30.3%) and culture (31.4%), indicating a higher sensitivity of the present method than the traditional ones. The feasibility of this method was further approved by successful detection of Neisseria gonorrhoeae and Chlamydia trachomatis.

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