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Colloidal gold immunochromatography test paper for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin compound and preparation method

An immunochromatographic detection and globin-binding technology, applied in the field of immunodiagnosis, can solve problems such as inaccurate detection results, and achieve the effects of direct detection results, intuitive display, and easy operation.

Pending Publication Date: 2020-11-03
HANGZHOU ALLTEST BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The present invention aims to overcome the problem in the prior art that the hemoglobin in the digestive tract is easily destroyed by pepsin, resulting in inaccurate detection results, and provides a colloidal gold immunochromatographic detection test paper and preparation method for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin complexes. The joint detection of hemoglobin + hemoglobin-hemoglobin complex improves the sensitivity of occult blood detection including upper gastrointestinal bleeding, and the two-in-one detection reagent of hemoglobin and hemoglobin-hemoglobin complex can simultaneously detect upper and lower gastrointestinal bleeding

Method used

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  • Colloidal gold immunochromatography test paper for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin compound and preparation method
  • Colloidal gold immunochromatography test paper for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin compound and preparation method
  • Colloidal gold immunochromatography test paper for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin compound and preparation method

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preparation example Construction

[0046] Preparation of mouse anti-human hemoglobin monoclonal antibody: (1) BALB / C mice eight weeks old were immunized with antigens, injected subcutaneously, injected once every three weeks, immunized five times in total, blood was collected to measure antibody titer by ELISA, The titer is good, and the injection is boosted three days before refusion; (2) Cell fusion and culture: the mice with super-immune 3d, the mice were killed by neck dislocation, the spleen was removed by aseptic surgery to prepare spleen cells, and SP2 / 0 cells were mixed with SP2 / 0 cells in PEG Under the effect of cell fusion, add the fused cell suspension to the cell culture plate, and place it in a 37°C, 5% CO 2 incubator for selective culture with HAT and HT medium; (3) Cell screening: use indirect competition E The positive wells were screened by LISA, and the wells with strong positive and vigorous cell growth were cloned by limiting dilution three times; (4) Production of monoclonal antibody in asci...

Embodiment 1

[0051] The preparation method of the colloidal gold immunochromatography test paper for rapid detection of human hemoglobin and haptoglobin-hemoglobin complexes specifically comprises the following steps:

[0052] 1) Prepare the first detection line T and the first quality control line C: dilute the mouse anti-human hemoglobin monoclonal antibody and spray it on the nitrocellulose membrane at a speed of 1.0-1.2ul / cm as the first detection line T, Goat anti-mouse IgG antibody solution was diluted and sprayed on the nitrocellulose membrane as the first quality control line C, and dried at 36°C to obtain the detection line and quality control line; the detection line T coated mouse anti-human The concentration of hemoglobin monoclonal antibody is 0.5mg / ml, and the concentration of goat anti-mouse IgG antibody is 1.0mg / ml;

[0053] 2) Preparation of the second detection line T and the second quality control line C: Dilute the mouse anti-human haptoglobin-hemoglobin complex monoclo...

Embodiment 2

[0057] The preparation method of the colloidal gold immunochromatography test paper for rapid detection of human hemoglobin and haptoglobin-hemoglobin complexes specifically comprises the following steps:

[0058] 1) Prepare the first detection line T and the first quality control line C: Dilute the mouse anti-human hemoglobin monoclonal antibody and spray it on the nitrocellulose membrane at a speed of 1.15ul / cm as the first detection line T, goat antibody Mouse IgG antibody solution was diluted and sprayed on the nitrocellulose membrane as the first quality control line C, and dried at 36.5°C to obtain the test line and quality control line; the test line T coated mouse anti-human hemoglobin single Cloning antibody concentration is 0.8mg / ml, goat anti-mouse IgG antibody concentration is 1.5mg / ml;

[0059] 2) Prepare the second detection line T and the second quality control line C: Dilute the mouse anti-human haptoglobin-hemoglobin complex monoclonal antibody and spray it on t...

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Abstract

The invention relates to the technical field of immunodiagnosis, and discloses colloidal gold immunochromatography test paper for rapidly diagnosing hemoglobin and haptoglobin hemoglobin complex and apreparation method thereof, aiming at the problem of inaccurate detection result caused by easy damage of pepsase to hemoglobin in digestive tracts in the prior art. The method specifically comprisesthe following steps: 1) preparing a first detection line T and a first quality control line C; 2) preparing a second detection line T and a second quality control line C; 3) preparing a gold-labeledpad; 4) preparing the test paper: sequentially connecting and assembling the water absorption pad, the sample pad, the microsphere probe treatment pad, the antibody-coated modified nitrocellulose membrane and the PVC bottom plate to obtain a finished product. The hemoglobin and hemoglobin combined globin compound is adopted for combined detection, the detection sensitivity of occult blood including upper gastrointestinal hemorrhage is improved, operation is easy, convenient and rapid, the result is visually and accurately displayed, sensitivity is high, specificity is good, and the preparationprocess is simple.

Description

technical field [0001] The invention relates to the technical field of immunodiagnosis, in particular to a colloidal gold immunochromatographic detection test paper for rapidly diagnosing hemoglobin and haptoglobin-hemoglobin complexes and a preparation method thereof. Background technique [0002] Hemoglobin (Hb, Hemoglobin) in feces, also known as fecal occult blood (FOB, Fecal Occult Blood), is currently clinically regarded as one of the tumor markers for the diagnosis of rectal cancer and colon cancer. Hemoglobin easily combines with haptoglobin to form a stable and irreversible complex called hemoglobin-haptoglobin complex (Hb-Hp Complex). Anyone with a small amount of bleeding caused by gastrointestinal diseases may have occult blood in the stool. Therefore, occult blood examination is of great value in diagnosing a variety of gastrointestinal bleeding diseases. It is an effective means for general screening of gastrointestinal diseases. research hotspot. [0003] Im...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/558G01N33/577
CPCG01N33/558G01N33/577
Inventor 陈金树陆维克涂小娟高飞
Owner HANGZHOU ALLTEST BIOTECH
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