No abstract

Microtiter plate-format optodes could be assembled by casting bulk-response membranes into the standard 96-well polypropylene-based plate or by screen printing them on an optically transparent substrate with 96-well pattern. The compositions of thick optode membranes, especially the ratios of poly(vinyl chloride) (PVC) to plasticizer [bis-(2-ethylhexyl) sebacate (DOS)], were carefully optimized to provide reproducible and rapid response. Adjusting the ratio of PVC to DOS by 1:6, bulk-response membranes containing neutral carrier (4-tert-butyl calix[4]arene tetraacetic acid tetraethyl ester for sodium-selective membrane or valinomycin for potassium-selective membrane) and lipophilic pH indicator (ETH 5294) could exhibit equilibrium response in 5 min. The practical utility of microtiter plate-format optodes has been examined by determining clinically relevant electrolytes in serum samples. It was demonstrated that microtiter plate-format optodes can provide high sample throughput (approximately 100 samples in less than 5 min), analytical performance comparable to that of a potentiometric clinical analyzer, and additional information on electrolytes using the same samples prepared for other colorimetric measurements.

The strong interaction between biotin and avidin is utilized for the development of enzyme-linked competitive binding assays not only for biotin itself, but also for other biomolecules. Alkaline phosphatase, a single substrate enzyme typically used for heterogeneous types of competitive binding assays, is employed also for homogeneous types. For the biotin assay, the heterogeneous protocol offers a much improved detection capability when compared to the homogeneous type. A simple approach of simulating dose-response curves is introduced. An effective analyte concentration attached to an enzyme label is determined by using the same binder, but with a different enzyme label. The analyte system adapted to the biotin/avidin-mediated homogeneous protocol is digoxin and a monoclonal anti-digoxin antibody. The detection capabilities of these assays, particularly the homogeneous types, are shown to be limited by the detectability of the enzyme conjugate employed.

Various types of synthetic polymers are widely used to immobilize or entrap natural and artificial receptors to impact a molecular-recognition capability to a sensitive transducer of chemical sensors.1,2 A poly(vinyl chloride) (PVC)-based ionselective membrane is a typical example: it is readily prepared by incorporating small amounts of electroactive compounds (1 -2% of ionophore and lipophilic salts) into a solvent polymeric matrix comprising one part high molecular-weight PVC and two parts plasticizer.3 Also, many researchers have been employing alternative polymer matices (e.g. silicone rubber (SR) and polyurethane (PU)), because of some inherent problem associated with the use of PVC-based membranes. Each matrix possesses its own advantages and disadvantages. 4 Recently, our group reported that the membranes are formulated with either RTV-SR or aromatic-type PU as the matrices, and that those optimized membranes (i.e. incorporated both cationic and anionic-exchange sites and plasticizer) could be applied as a new type of liquid junction-free reference electrode system. 5-7 Particularly, the aromatic PU matrix formulation exhibits only a small emf variation, even at very high salt concentrations and no appreciable response over a wide pH range. However, the SR-based membrane, while possessing minimal response toward most ions, is found to display a near-Nernstian response to pH change. It is a very interesting phenomenom that a polymer membrane fabricated without proven H + ionophore, such as tridodecylamine (TDDA), exhibits a near-Nernstian response to H + ion over a wide pH range. Even the potentiometric behavior of an RTV-SR membrane doped with TDDA is non-Nernstian, exhibiting an inflection point at pH 6.4 due to a drastic change in its response slope from 94 to 40 mV/pH. 8 However, the calibration plots for the RTV-SR/TDDA membranes are gradually rectified as the content of the plasticizer is increased. The cause of those aforementioned non-ideal behaviors is not yet fully understood. Thus, we examined three different classes (i.e. PVC and two types of RTV-SR) of solvent-processible polymeric membranes at various incorporating compositions with anionic and cationic ion-exchangers for the possibility of pH measurements. Experimental ReagentsThe sources of the reagent used were as follows: high molecular-weight PVC, bis(2-ethylhexyl)adipate (DOA), onitrophenyl octyl ether (NPOE), tridodecylmethylammonium chloride (TDMACl), potassium tetrakis(p-chlorophenyl)borate (KTpClPB) and tetrakis[tris(dimethylamino)phoranylidenamino]-phosphorium chloride (TPAPCl) from Fluka (Buch, Switzerland); and 3140 RTV-SR and 730 RTV-SR from Dow Corning (Midland, MI). All other chemicals used were of analytical reagent grade. Standard solutions and buffers were prepared with deionized water. Preparation and evaluation of polymer membranes and electrodesPolymer membranes were prepared according to a previously reported method. 9 The detail compositions of the optimized membranes used in this experiment are listed in Table 1. The...