1. Field of the Invention
This invention relates to methods, compositions and kits for detecting hydrogen peroxide or a compound capable of generating hydrogen peroxide.
The clinical diagnostic field has seen a broad expansion in recent years, both as to the variety of materials of interest that may be readily and accurately determined, as well as the methods for the determination. Convenient, reliable and non-hazardous means for detecting the presence of low concentrations of materials in liquids is desired. In clinical chemistry these materials may be present in body fluids in concentrations below 10.sup.-12 molar. The difficulty of detecting low concentrations of these materials is enhanced by the relatively small sample sizes that can be utilized.
Detection of low concentrations of hydrogen peroxide is useful for numerous analytical procedures, particularly in clinical chemistry. Hydrogen peroxide is produced by cells such as monocytes and is an indicator of monocyte activation. Additionally, any material of interest that is or can be converted to a substrate for an oxidase such as xanthene oxidase, amino acid oxidase, NADH oxidase, glucose oxidase, galactose oxidase, glycerol phosphate oxidase, and the like can be detected by the hydrogen peroxide that is produced by the action of the enzyme on the substrate. Tests for glucose, triglycerides, d-amino acids, and cholesterol can be routinely carried out by detecting hydrogen peroxide, usually by reaction of a peroxidase and a chromogenic substrate. Enzyme immunoassays using an oxidase such as glucose oxidase as a label also depend on a sensitive detection method for hydrogen peroxide. For example, when glucose oxidase is used as a label, the hydrogen peroxide can be detected using horseradish peroxidase and a chromogenic substrate, or the hydrogen peroxide can be detected electrochemically.
Detection of hydrogen peroxide is also becoming more important in the area of foodstuffs. For example, in some countries hydrogen peroxide is used as a bleaching agent for food. It is important that residual levels of hydrogen peroxide in the food after bleaching be substantially zero to avoid health hazards.
A method that has higher sensitivity, less interference from the sample, and uses fewer, and more stable, reagents would increase the simplicity and reliability of assays for, or depending on, hydrogen peroxide detection.
Homogeneous immunoassays have previously been described for small molecules. These assays include Syva Company's FRAT.RTM. assay, EMIT.RTM. assay, enzyme channeling immunoassay, and fluorescence energy transfer immunoassay (FETI); enzyme inhibitor immunoassays (Hoffman LaRoche and Abbott Laboratories): fluorescence polarization immunoassay (Dandlicker), among others. All of these methods have limited sensitivity, and only a few including FETI and enzyme channeling, are suitable for large multiepitopic analytes.
Luminescent compounds, such as fluorescent compounds and chemiluminescent compounds, find wide application in the assay field because of their ability to emit light. For this reason, luminescers have been utilized as labels in assays such as nucleic acid assays and immunoassays. For example, a member of a specific binding pair is conjugated to a luminescer and various protocols are employed. The luminescer conjugate can be partitioned between a solid phase and a liquid phase in relation to the amount of analyte in a sample suspected of containing the analyte. By measuring the luminescence of either of the phases, one can relate the level of luminescence observed to a concentration of the analyte in the sample.
Particles, such as liposomes and erythrocyte ghosts, have been utilized as carriers of encapsulated water soluble materials. For example, liposomes have been employed to encapsulate biologically active material for a variety of uses, such as drug delivery systems wherein a medicament is entrapped during liposome preparation and then administered to the patient to be treated.
Particles, such as latex beads and liposomes, have also been utilized in assays. For example, in homogeneous assays an enzyme may be entrapped in the aqueous phase of a liposome labeled with an antibody or antigen. The liposomes are caused to release the enzyme in the presence of a sample and complement. Antibody or antigen-labeled liposomes, having water soluble fluorescent or non-fluorescent dyes encapsulated within an aqueous phase vesicle or lipid soluble dyes dissolved in the lipid bilayer of a lipid, have also been utilized to assay for analytes capable of entering into an immunochemical reaction with the surface bound antibody or antigen. Detergents have been used to release the dyes from the aqueous phase of the liposomes.
2. Brief Description of the Related Art
U.S. Pat. No. 5,084,381 (Akimoto, et al.) discusses an assay method for detecting hydrogen peroxide.
Processes and materials for carrying out specific binding assays is disclosed in patent application WO 86/01899 (Davis, et al.).
U.S. Pat. No. 5,108,893 (Baret) discloses the use of oxidase enzyme systems in chemiluminescent assays.
European Patent Application 0 421 788 A2 (Allen) discloses a haloperoxidase-acid-oxidant chemiluminescence assay system for determining the presence or amount of an analyte in a liquid sample.
U.S. Pat. No. 4,315,998 discusses polymer-bound photosensitizing catalysts.
Photoactivatable chemiluminescent matrices are described in patent application WO 94/03812 (Pease, et al.).
European Patent Application No. 0 515 194 A2 discloses assay methods utilizing induced luminescence. The references cited therein are incorporated herein by reference including without limitation U.S. Pat. No. 5,017,473 (Wagner), which discloses a homogeneous chemiluminescence immunoassay using a light absorbing material, European Patent Application No. 0,345,776 (McCapra), which discloses specific binding assays that utilize a sensitizer as a label, U.S. Pat. No. 4,193,983 (Ullman, et al.), which discloses labeled liposome particle compositions and immunoassays therewith, U.S. Pat. No. 4,891,324 (Pease, et al.), which describes a particle with luminescer for assays.