Method for quantitative detection of biological toxins

• US 20070172904A1
• Filed: 11/24/2004
• Published: 07/26/2007
• Est. Priority Date: 11/24/2004
• Status: Abandoned Application

• Current Assignee: Institut Bioorganicheskoi Khimii, Institut Molekulyarnoi Biologii IMVA Engelgardta Rossiiskoi Akademii NAUK
• Original Assignee: Institut Bioorganicheskoi Khimii, Institut Molekulyarnoi Biologii IMVA Engelgardta Rossiiskoi Akademii NAUK
• Inventors: Dementieva, Ekaterina, Stomakhin, Andrei, Rubina, Alla, Grishin, Evgeny, Djukova, Veronika, Nesmeyanov, Vladimir, Zasedatelev, Alexander
• Application Number:
  US10/589,143
• Publication Number:
  US 20070172904A1
• Time in Patent Office: Days
• US Class Current: 435/7.320
• CPC Class Codes: B01J 2219/00497 Features relating to the solid phase supports
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00576 fluorophore
• CPC Class Codes: B01J 2219/00578 electrophoric
• CPC Class Codes: B01J 2219/00581 Mass
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00644 the porous medium being present in discrete locations, e.g. gel pads
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00677 Ex-situ synthesis followed by deposition on the substrate
• CPC Class Codes: B01J 2219/00691 using robots
• CPC Class Codes: B01J 2219/00711 Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B82Y 30/00 Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: G01N 33/5308 for analytes not provided for elsewhere, e.g. nucleic acids, uric acid, worms, mites
• CPC Class Codes: G01N 33/56911 Bacteria

Abstract

The invention relates to analytical chemistry and to quantitative immunochemical analysis, in particular, to a method for immunochemical quantitative detection of various biological toxins by using biological microchips. A biological microchip comprises an ordered array of three-dimensional hydrogel cells on a solid support, which are produced by a method of photo- or chemically induced polymerization and contain immobilized antibodies to various bacterial, plant and animal biotoxins, or biotoxins. The use of microchips makes it possible to analyze a sample simultaneously for the presence of several biotoxins. The proposed method for detecting biotoxins can be used in medicine, in food industry, and in environmental protection.

Claims

1. A method for quantitative detection of biotoxins in a sample, comprising the steps of:

a) manufacturing a biological microchip comprising an ordered array of three-dimensional hydrogel elements on a solid support, obtained by a method of photo- or chemically induced polymerization and containing immobilized antibodies to various bacterial, plant or animal toxins or biotoxins, wherein an antibody to an individual biotoxin or an individual biotoxin is immobilized in each separate cell,

b) incubating the microchip in a reaction medium which comprises a sample containing biotoxins to be analyzed, for forming immune biotoxin-antibody complexes, which incubation, when necessary, is carried out under stirring conditions;

c) detecting the formed complex;

d) quantitative detection of the biotoxin being analyzed.

2. The method as claimed in claim 1, wherein the immobilized antibodies comprise antibodies selected from the group of antibodies to ricin, viscumin, staphylococcal entero-toxin B, tetanus toxin, diphtheria toxin, lethal factor of anthrax toxin.

3. The method as claimed in claim 1, wherein the immobilized biotoxins comprise biotoxins selected from the group comprising ricin, viscumin, staphylococcal entero-toxin B, tetanus toxin, diphtheria toxin, lethal factor of anthrax toxin.

4. The method as claimed in claim 1, wherein detecting the complex formed in step c) and subsequent quantitative detection in step d) are carried out in a format of direct immunoassay.

5. The method as claimed in claim 1, wherein the reaction medium in step b) additionally contains antibodies to a biotoxin, and the detection of the complex formed between the biotoxin immobilized on the chip and the antibody against this biotoxin in step c) and subsequent quantitative detection in step d) are carried out in the format of competitive immunoassay.

6. The method as claimed in claim 1, wherein the reaction medium in step b) further contains a labeled biotoxin, and detection of the complex formed between the antibody immobilized on the chip and the biotoxin in step c) and subsequent quantitative detection in step d) are carried out in the format of competitive immunoassay.

7. The method as claimed in claim 1, wherein detection the complex formed in step c) and subsequent quantitative detection in step d) are carried out in the format of sandwich-immunoassay.

8. The method as claimed in claim 1, wherein quantitative detection of the biotoxin is effected by carrying out steps a)-c) with known concentrations of the biotoxin being analyzed and with plotting a calibration dependence curve, from which the amount of the biotoxin being analyzed in the sample is determined.

9. The method as claimed in claim 1, wherein in step c) detection of the formed complex is carried out fluorimetrically, chemiluminometrically or mass-spectrometrically.