Search

In order to the increase sensitivity of enzyme-linked immunosorbent assay (ELISA) using micro-droplet reaction system formed by ink-jet, we employed small dots of two-dimensionally well-ordered structure of polystyrene (PS) microbeads (710 +/- 25 microm in diameter) formed on the surface of polydimethyl siloxane (PDMS). An aqueous suspension of polystyrene microbeads (5 microm) was ejected on the PDMS plate with ink-jet. The PS beads were automatically assembled by capillary force accompanied with solvent evaporation. The evaporation rate was controlled by ambient relative humidity. The assembled beads were solidly immobilized on the surface of PDMS surface. The dots of well-ordered structure were stable against vigorous washing. ELISA using the structure as a reaction location was three times more sensitive than the material without the structure due to the increase of the surface area and consequent increase of the amount of antibody.

An ultra-micro sample injector for gas chromatography (GC) was developed. An ink-jet microchip, originally used for industrial recorder, was modified at the edge near to an orifice, and fixed into the GC. In order to evaluate the characteristics of this injector, a sample injector and a thermal conductive detector (TCD) were connected directly, while water was used as the test sample. The volume of the droplet, the interval time and the back-pressure to the ink-jet microchip were investigated. Within the range of 1 - 5 nL volume injected sample, the TCD response according to the amount of the sample volume (the volume of one droplet from the ink-jet microchip was about 1 nL) was obtained. A good reproducibility of the peak area was obtained to be about 1.0% of the RSD value. In order to compare the injection method of the ink-jet chip with that using a micro-syringe, the method using the ink-jet chip could introduce 1/1000 of the amount of the sample and gave reproducible results.

A surface-reaction system in a nanoliter water pool using an ink-jet microchip was developed. The reaction system in the nanodroplets formed on a poly(dimethylsiloxane) (PDMS) coated glass slide increased the diffusion-controlled reaction without using a nano-pump, specialized connector or highly sensitive detector. When nanoliter droplets were placed on the PDMS surface with a distance of 100 microm between them by the ink-jet microchip, the repeatabilities of the fluorescence intensity were 2.9% RSD (n = 7). The used ink-jet microchip had 4 different injection ports, and the distance between the ports was 0.995 mm. It was necessary to correct the distance in order to mix or dilute samples in a small droplet. The correction was successfully performed by moving the X-Y stage using inhouse-made software. A linear relationship was obtained between the Resorufin concentrations and the fluorescence intensity. We applied this system to an enzyme-linked immunosorbent assay (ELISA) for immunoglobulin A (IgA), and observed a difference in the fluorescence intensity derived from the amount of IgA (blank, 6.25 ng/mL, 12.5 ng/mL). These results show the usefulness of the open-type micro-analytical systems proposed by us.

The chiral separation of amino acid derivatives by ligand-exchange electrophoresis in a microchannel chip was performed for the first time. A Cu(II) complex with L-prolinamide was used as a chiral selector. The migration behaviors of eleven NBD-DL-amino acids were investigated by ligand-exchange capillary electrophoresis (LE-CE). The enantiomer of five NBD-amino acids (Ser, Thr, Val, Phe and His) could be separated by LE-CE using a 20 mM ammonium acetate buffer (pH 9.0) containing 10 mM copper acetate, 20 mM L-prolinamide and 1 mM SDS. NBD-His was eluted in the order D-form and L-form, while the elution order of another enantiomers was L-form and D-form. Under this condition, the enantioseparation of these five NBD-amino acids by ligand-exchange microchip electrophoresis (LE-ME) was investigated using a glass microchip. The enantioseparation of NBD-Ser, -Thr and -His could be successfully accomplished by LE-ME. LE-ME was superior to LE-CE in terms of the short migration time and a good enantiomeric separation.

The photo-responses of the retention and enantioseparation of several optical isomers were evaluated using an azobenzene-modified gamma-cyclodextrin stationary phase (Az gamma-CDSP) in micro-HPLC. UV light irradiation induced a decrease in the retention and the chiral selectivity for N-(3,5-dinitrobenzoyl)-1-phenylethylamine (DNBPEA) and N-(3,5-dinitrobenzoyl)-1-(1-naphtylethyl)amine (DNBNEA), while an increase was induced for dansylphenylalanine (DnsPhe) using a mixture of methanol and aqueous phosphate buffer as the mobile phase. No changes in the retention and the enantiomer separation of benzoin were observed with UV light irradiation. The retention behaviors were recovered by visible-light irradiation. It was speculated that the main factor of the change in the retention behavior was a change in the pi-pi interaction due to the azobenzene moiety of the stationary phase with photo-irradiation. Comparing the retention behavior before and after UV light irradiation, a suitable condition for obtaining a better resolution and enantiomer separation would be chosen using Az gamma-CDSP.