CPC G01N 33/54346 (2013.01) [G01N 33/545 (2013.01); G01N 33/68 (2013.01); G01N 2333/4727 (2013.01)] | 10 Claims |
1. A method for measuring the presence of calprotectin in a biological sample of a patient by a particle-enhanced turbidimetric immunoassay using a test kit comprising:
a first aqueous reagent-component comprising:
20 to 1000 mmol/L organic buffer with a pH in the range from 5.0 to 6.0;
50 to 300 mmol/L salt of sodium, potassium or lithium;
0.1 to 1.5% protease-treated serum albumin;
0.01 to 0.1% (w/v) sodium dodecyl sulfate, and
optionally beta-aldoses, triose, tetroses, pentoses, hexoses, glucan, dextran and/or sugar to achieve an osmolality of at least 200 mosmol/kg of H2O; and
a second reagent-component comprising:
0.01 to 0.5% (w/v) latex particles of 150 to 350 nm diameter carrying immobilized monoclonal antibodies which bind either one of S100A8 and S100A9 proteins or calprotectin (S100A8/A9);
the method comprising the steps of:
a) collecting a predetermined amount of said biological sample;
b) solubilizing and extracting said biological sample in the first aqueous reagent-component having i) a pH between 5.0 and 6.0, ii) an osmolality of at least 150 mosmol/kg of H2O, iii) wherein the organic buffer is co-coordinating and sequestering calcium and zinc ions, and optionally, homogenizing and extracting the matrix of said biological sample followed by a removal of any particulate material to obtain a sample solution with a solubilized presence of essentially heterodimeric calprotectin (S100A8/A91);
c) mixing a defined amount of said sample solution of step (b) with an amount of the second reagent-component comprising latex nanoparticles having immobilized two or more monoclonal antibodies or fragments thereof which specifically bind either one of S100A8 and S100A9 or calprotectin (S100A8/A9) to obtain a particle-bound antibody-antigen reaction with calprotectin (S100A8/A9) being present in a defined molecular state;
d) incubating the mixture of step c) for an interval of time; and
e) acquiring an optical property of the mixture and determining a signal indicative of the content of calprotectin (S100A8/A9) based on the optical property of the mixture;
f) relating said content to a calibrated control and assessing the clinical condition of said patient based on the measured presence of calprotectin (S100A8/A9) in said biological sample.
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