Electrochemiluminescent assay

J.G. Bruno and J.C. Cornette. Electrochemiluminescence Assays Based on Interactions with Soluble Metal Ions and Diaminoaromatic Ligands, US patent no. 5 976 887. 1999. Available at http //www.freepatentsonline.com/5976887.html. 

The availability of ECL technology from instrument providers such as Meso Scale Discovery (MSD) has enabled practical applications of electrochemiluminescent assays. The MSD platform uses carbon electrode plates that are available uncoated or coupled with biotin or streptavidin. Thus, the platform may be used with direct, indirect, and bridging formats. 

Kenten, J.H., Casadei, J., Link, J., Lupoid, S., Willey, J., Powell, M., Rees, A., and Massey,R. (1991) Rapid electrochemiluminescence assays of polymerase chain reaction products. Clinical Chemistry, 37, 1626 1632. 

FIGURE 24.1 BNP concentrations in rat plasma. BNP concentrations in plasma from rats administered rosiglitazone (80mg/kg) by oral gavage for 4 days (n= 10). BNP was measured using Meso Scale Discovery s rat BNP enzyme-linked electrochemiluminescence assay 4h after the fourth daily dose. 

Zhou M, Roovers J, Robertson GP, Grover CP (2003) Multilabeling biomolecules at a single site. 1. Synthesis and characterization of a dendritic label for electrochemiluminescence assays. Anal Chem 75(23) 6708-6717... 

Weinreb PH, Yang WJ, Violette SM, Couture M, Kimball K, Pepinsky RB, Lobb RR, Josiah S (2002) A cell-free electrochemiluminescence assay for measuring betal-integrin-ligand interactions. Anal Biochem 15 306(2) 305-313... 

In this paper the electtode anodic reactions of a number of dihydropyridine (DHP) derivatives, quantum-chemical calculations of reactions between DHP cation-radicals and electrochemiluminescers anion-radicals (aromatic compounds) and DHP indirect ECL assay were investigated. The actuality of this work and its analytical value follow from the fact that objects of investigation - DHP derivatives - have pronounced importance due to its phaiTnacology properties as high effective hypertensive medical product. 

Probably the most sensitive biosensor for rapid screening has been the ORIGIN electrochemiluminescence system (IGEN, Gaithersburg, MD). The primary reason for the ORIGIN S enhanced sensitivity is that it preconcentrates the target prior to the assay. Immunomagnetic beads are... 

Kuczynska E., Boyer D.G., Shelton D.R., Comparison of immunofluorescence assay and immunomagnetic electrochemiluminescence in detection of Cryptosporidium parvum oocysts in karst water samples, J. Microbiol. Meth. 2003 53 17-26. 

The model immunoassay is the enzyme-linked immunosorbent assay (ELISA) in which a non-specific capture antibody is bound to a surface, such as a multi-well plate or small tube [13]. In the basic form of ELISA, a second antibody tagged with an enzyme interacts specifically with the analyte. The enzyme assay produces a colored product that is read with a spectrophotometer. There are many variations on the basic immunoassay format that serve to increase sensitivity, specificity, linear range, and speed. Many commercial instruments have been developed to take advantage of various technologies for reporter molecules. The immunoassay may be coupled to an electronic sensor and transducer, such as a surface acoustical wave (SAW) sensor. Electrochemiluminescence (ECL) is a method in which the detector antibody is tagged with a ruthenium-containing chelate [13-15]. When the tag is... 

Assay for human immunodeficiency virus type 1 (HIV-1) proviral DNA in peripheral blood monuclear cells can be performed by PCR followed by detection of PCR products by electrochemiluminescence-labeled oligonucleotide probe [Tris-bipyridine ruthenium (II) complex]. Since one of the PCR primers is biotin-labeled at the 5 end, facile capture of the PCR product-probe complex can be accomplished on streptavidin-conjugated magnetic particles, prior to analysis in an electrochemiluminescence analyzer (S3). 

On the other hand, the main types of immunoassays that can be performed by using labelled antibodies or antigens are direct sandwich, competitive and indirect assays. The labels can be enzymes (alkaline phosphatase, peroxidise or glucose oxidase) metal NPs (gold) fluorescent or electrochemiluminescent probes. 

Immunogenicity assays for investigating the frequency and consequences of antibody development against a protein therapeutic agent are typically based on an immunoassay technique (mostly ELIS As of various types). However, other assay formats are available such as radioimmunoprecipitation assay, surface plasmon resonance, and electrochemiluminescence [3]. Assays for measuring antibody response should be established in the early preclinical stage of development to estimate the value of the applied animal models (see Chapters 16 and 20). 

Chemiluminescence immunoassay These assays rely on the use of chemiluminescent labels, which may be labelled antigens or antibodies. For example, the chemiluminescent label isoluminol is oxidized in the presence of hydrogen peroxide and a catalyst, producing long-lived light emission that is measured, thus allowing determination of the unknown antigen or antibody concentration in a sample. Another useful variant is electrochemiluminescence immunoassay that is commonly used in biomolecular detection, and in particular the measurement of native and recombinant peptides and proteins. 

Blackburn GF, Shah HP, Kenten JH, et al. Electrochemiluminescence development of immunoassays and DNA probe assays for clinical diagnostics. Clin Chem 1991 37 1534-9. 

Ruthenium (II) tris(bipyridyl) (Figure 9-17, B) undergoes an electrochemiluminescent reaction (620 nm) with tripropylamine at an electrode surface, and this chelate is now used as a label in competitive and sandwich electrochemiluminescence immunoassays. Using this label, various assays have been developed in a flow cell using magnetic beads as the solid phase. Beads are captured at the electrode surface, and an unbound label is washed out of the cell by a wash buffer. Label bound to the bead undergoes an electrochemiluminescent reaction, and the light emission is measured by an adjacent photomultiplier tube. ... 

For each type of assay, the label or tag involved is listed along with typical substrates, type of absorbance, and type of instrument required. RIA radioimmunoassay IRMA immunoradiometric assay CPM counts per minute ELISA enzyme linked immunosorbent assay EIA enzyme immunoassay TMB 5,5 tetramethylbenzidine HPA p hydroxyphenylacetic acid AMPPD 4 methoxy 4 (3 phosphatephenyl) spiro(l,2 dioxetane) 3,2 adamantane p NPP p nitrophenyl phosphate 4 MUP 4 methylumbelliferyl phosphate ONPG o nitrophenyl P galactopyranoside MUG 4 methylumbelliferyl P D galactopyrano side AMPGD 3 (4 methoxyspiro( 1,2 dioxetane 3,2 tricyclo(3.3.1.1(3,7))decan) 4 yl)phenylgalacto pyranoside ECL electrochemiluminescence. 

---