J U Eskola’s research while affiliated with Aalto University and other places

Electrochemistry of hot electrons in fully aqueous solutions at tetrahedral amorphous carbon thin film electrodes is discussed. The generation of these highly reducing chemical species was confirmed by normal pulse voltammetry and several electrochemiluminescent systems. Electron transfer into pre‐existing solvent cavities was observed at approximately ‐2.65 V vs. Ag/AgCl (sat.). Electrogenerated hot electrons were utilized as chemiluminescent mediators in heterogeneous sandwich immunoassay of Serum Amyloid A. The calibration curve was linear over four orders of magnitude and the detection limit was 85 ng L‐1 that demonstrates the efficiency of hot electron generation at this electrode material.

Novel hot electron-emitting working electrodes and conventional counter electrodes were created by screen printing. Thus, low-cost disposable electrode chips for bioaffinity assays were produced to replace our older expensive electrode chips manufactured by manufacturing techniques of electronics from silicon or on glass chips. The present chips were created by printing as follows: (i) silver lines provided the electronic contacts, counter electrode and the bottom of the working electrode and counter electrode, (ii) the composite layer was printed on appropriate parts of the silver layer, and (iii) finally a hydrophobic ring was added to produce the electrochemical cell boundaries. The applicability of these electrode chips in bioaffinity assays was demonstrated by an immunoassay of human C-reactive protein (i) using Tb(III) chelate label displaying long-lived hot electron-induced electrochemiluminescence (HECL) and (ii) now for the first time fluorescein isothiocyanate (FITC) was utilized as an a low-cost organic label displaying a short-lived HECL in a real-world bioaffinity assay.

Aromatic Tb(III) chelates can be detected down to subnanomolar concentrations at polystyrene-carbon black composite electrodes on the basis of hot electron-induced electrochemiluminescence, and used as electrochemiluminescent labels in bioaffinity assays. The excitation mechanism is based on chemiluminescent reactions that are initiated by the field emission/tunnel emission of hot electrons from the composite electrode. The composition and the properties of the novel composite electrodes were studied in detail. Hot electron-induced electrochemiluminescence intensities obtainable with the present composite electrodes are comparable to the previously used metal/insulator-type electrodes without exhibiting considerable long-lived solid state electroluminescence background emission, unlike e.g. oxide-coated silicon and aluminum electrodes. C-reactive protein was finally determined in a heterogeneous sandwich-assay by using the present composite electrodes as a solid support material for the capture antibodies. The detection antibody was labeled with a commercially available Tb(II) chelate. Calibration curve for the determination of C-reactive protein was linear over two orders of magnitude and the detection limit was well below the clinical reference value, approx. 1 μg l⁻¹. Disposable polystyrene-carbon black composite electrodes are superior alternatives for e.g. silicon-based electrodes due to the low cost, easier manufacturing process and a very good performance e.g. in immunoassays.

A homogeneous immunoassay of T4 was developed using a semiautomatic electrochemiluminometer modified from a commercially available fluorometer. In addition, from the same analyte panel an immunometric immunoassay of TSH at similar disposable oxide-covered aluminum rake electrodes was studied using this instrument both on homogeneous and heterogeneous basis. Detection was based on hot electron-induced cathodic electrochemiluminescence utilizing a commercially available Tb(III) chelate label. The assays were reasonably sensitive and comparison was made with other older methods. Thus, it is possible to develop both non-competitive and competitive immunoassays based on detection of hot electron-induced ECL of the labels.

Double insulating barrier tunnel emission electrodes were fabricated by adding a new pure aluminum layer upon oxidized aluminum electrodes by vacuum evaporation and thermally oxidizing the new aluminum layer in air at room temperature. Resulting Al/Al2O3/Al/Al2O3 electrodes allow the use of various aluminum alloys in the electrode body necessary for hardness or shaping ability of the electrode while obtaining the luminescence properties of pure aluminum oxide. During electrical excitation of luminescent labels by cathodic hot electron injection into aqueous electrolyte solution, the background noise is mainly based on high-field-induced solid-state electroluminescence and F-center luminescence of the outer aluminum oxide film. The more defect states and/or impurity centers the outer oxide film contains, the higher is the background emission intensity. The present electrode fabrication method provides a considerable improvement in signal-to-noise ratio for time-resolved electrochemiluminescence (TR-ECL) measurements when the original native oxide film of the electrode body contains luminescence centers displaying long-lived luminescence. The excellent performance of the present electrodes is demonstrated by extremely low-level detection of Tb(III) chelates, luminol, Pt(II) coproporphyrin and Tb(III) labels in an immunometric immunoassay by time-resolved electrochemiluminescence.

Double insulating barrier tunnel emission electrodes were fabricated by adding a new pure aluminum layer upon oxidized aluminum electrodes by vacuum evaporation and thermally oxidizing the new aluminum layer in air at room temperature. Resulting Al/Al 2 O 3 /Al/Al 2 O 3 electrodes allow the use of various aluminum alloys in the electrode body necessary for hardness or shaping ability of the electrode while obtaining the luminescence properties of pure aluminum oxide. During electrical excitation of luminescent labels by cathodic hot electron injection into aqueous electrolyte solution, the background noise is mainly based on high-field-induced solid-state electroluminescence and F-center luminescence of the outer aluminum oxide film. The more defect states and/or impurity centers the outer oxide film contains, the higher is the background emission intensity. The present electrode fabrication method provides a considerable improvement in signal-to-noise ratio for time-resolved electrochemiluminescence (TR-ECL) measurements when the original native oxide film of the electrode body contains luminescence centers displaying long-lived luminescence. The excellent performance of the present electrodes is demonstrated by extremely low-level detection of Tb(III) chelates, luminol, Pt(II) coproporphyrin and Tb(III) labels in an immunometric immunoassay by time-resolved electrochemiluminescence.

Hot electrons can be injected from conductor/insulator/electrolyte (C/I/E) junctions into an aqueous electrolyte solution. Injected hot electrons induce electrogenerated chemiluminescence (ECL) of various luminophores in fully aqueous solutions. Such ECL gives the basis of electrochemiluminoimmunoassays and DNA-probe assays where different luminophores can be used as electrochemiluminescent labels. This work shows that SYBR® Green I is suitable as an ECL label in detection methods based on C/I/E tunnel-emission electrodes such as oxide-coated aluminium, magnesium and silicon electrodes.

Ruthenium(II) tris(2,2 -bipyridine) chelate shows chemiluminescence (CL) both during dissolution of metallic aluminium in alkaline conditions, and during dissolution of magnesium metal in acidic conditions. The presence of peroxodisulfate ions strongly enhances the CL. Magnesium system provides considerably better detectability of the present chelate giving linear calibration plot spanning over many orders of magnitude of concentration down to subnanomolar concentration levels. The possible primary species generated and luminescence mechanisms are shortly discussed.

Heterogeneous and homogeneous immunoassays of human thyroid stimulating hormone (hTSH) were developed on immunometric basis using aromatic Tb(III) chelates as electrochemiluminescent labels and varied types of disposable oxide-covered aluminum electrodes as the solid phase of the immunoassays. The long luminescence lifetime of the present labels allows the use of time-resolved electrochemiluminescence detection and provide the low detection limits of these labels and, thus, sensitive immunoassays. The primary antibody of immunometric immunoassays was coated upon aluminum oxide surface by physical absorption. In homogeneous immunoassays using 66 μl cell and 15 min incubation time, a linear calibration range of 0.25–324 μU/ml was obtained by applying only a single cathodic excitation pulse in the detection step of the assay.