Neutrophil gelatinase-associated lipocalin detection kit

A lipocalin and neutrophil technology, which can be used in biological tests, measuring devices, material inspection products, etc., and can solve problems such as inability to meet

Inactive Publication Date: 2018-04-17
NANTONG EGENS BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method takes several hours to detect NGAL, which cannot meet the cur

Method used

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Examples

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Effect test

Embodiment 1

[0035] The present embodiment provides a method for preparing an enzyme conjugate, comprising the following steps:

[0036] 1. Weigh an appropriate amount of Traut's reagent and prepare a 1.376 mg / mL solution with antibody activation buffer (100 mM triethanolamine buffer, pH 8.5).

[0037] 2. Weigh an appropriate amount of Sulfo-SMCC reagent and prepare a solution with a concentration of 17.5 mg / mL with DMF.

[0038] 3. Preserve the NGAL-labeled antibody in antibody activation buffer (100 mM triethanolamine buffer, pH 8.5) at a concentration of 2 mg / mL; add the Traut's solution prepared in step 1 to the solution, NGAL-labeled antibody and Traut' The molar ratio of s reagent is 1:15, mix immediately, and let stand at 25°C for 15 minutes.

[0039] 4. Add the Sulfo-SMCC solution prepared in step 2 to the alkaline phosphatase solution (AP solution, the concentration is 20mg / ml), the molar ratio of alkaline phosphatase and Sulfo-SMCC is 1:10, mix immediately, and let it stand at 2...

Embodiment 2

[0050] The present embodiment provides a kind of method for preparing enzyme conjugate, comprises the following steps:

[0051] 1. Weigh an appropriate amount of Traut's reagent, and use antibody activation buffer (100 mM triethanolamine buffer, pH 8.5) to prepare a 1.376 mg / mL solution.

[0052] 2. Weigh an appropriate amount of Sulfo-SMCC reagent, and use dimethylformamide DMF to prepare a solution with a concentration of 17.5 mg / mL.

[0053] 3. Store the NGAL-labeled antibody in antibody activation buffer (100 mM triethanolamine buffer, pH 8.5) at a concentration of 5 mg / mL; add the Traut's solution prepared in step 1 to the solution, NGAL-labeled antibody and Traut' The s reagent molar ratio is 1:30, mix immediately, and stand at 25°C for 15 minutes.

[0054] 4. Add the Sulfo-SMCC solution prepared in step 2 to the alkaline phosphatase solution (AP solution, the concentration is 20mg / ml), the molar ratio of alkaline phosphatase to Sulfo-SMCC is 1:15, mix immediately, and ...

Embodiment 3

[0065] This embodiment provides a method for magnetic bead coating, comprising the following steps

[0066] 1. Store the NGAL-coated antibody in 100mMMES buffer at pH 5.0 with a concentration of 1mg / mL;

[0067] 2. Take carboxyl magnetic bead solution 100 times the weight of the antibody, the particle size of carboxy magnetic beads is 3 μm, magnetically separate and discard the supernatant;

[0068] 3. Washing: re-dissolve the magnetic beads obtained in the above steps with 100 mMMES buffer solution of pH 5.0, and discard the supernatant by magnetic separation.

[0069] 4. Repeat step 3 once

[0070] 5. Add an appropriate amount of 100mMMES buffer to dissolve the magnetic beads obtained in step 4.

[0071] 6. Weigh an appropriate amount of NHS reagent and dissolve it in 100mMMES buffer with pH 5.0 to a solution with a concentration of 10mg / mL;

[0072] 7. Weigh an appropriate amount of EDC reagent and dissolve it in 100mMMES buffer with pH 5.0 to a solution with a concentra...

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Abstract

The invention discloses a neutrophil gelatinase-associated lipocalin detection kit which comprises seven storage assemblies, wherein the storage assemblies are respectively used for storing a neutrophil gelatinase-associated lipocalin (NGAL) calibration product, an NGAL quality control product, an enzyme conjugate working solution, a magnetic bead working solution, a cleaning solution, a substratesolution and a pretreatment reagent; the magnetic bead working solution contains carboxyl magnetic beads labeled with an NGAL antibody; and the magnetic bead working solution contains an anti-NGAL antibody labeled by alkaline phosphatase. The invention further discloses a detection method for detecting the neutrophil gelatinase-associated lipocalin. The detection kit disclosed by the invention has the lowest detection limit of 1ng/ml; and the linear range is 1-300ng/ml, the detection sensitivity is high, the linear range is wide, the detection duration is shortened to 15 minutes, and the detection steps are simplified.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to a neutrophil gelatinase-related lipocalin detection kit. Background technique [0002] Acute kidney injury (AKI) is a complex syndrome of renal insufficiency caused by a variety of reasons, which can occur in various clinical situations. It can manifest as a mild increase in serum creatinine levels or as acute renal failure. Studies have found that the occurrence of AKI is closely related to the increase in mortality. This requires physicians to make early diagnosis in clinical work. It is best to detect and intervene in time when the glomerular filtration rate begins to decline, or when there is only tissue damage and the glomerular filtration rate is still normal, so as to improve prognosis and reduce case fatality rate. Serum creatinine and urine output are the current diagnostic indicators of AKI, but they are affected by many factors, and their sensitivity and specific...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/543G01N33/535
CPCG01N33/6893G01N33/535G01N33/54326G01N2333/47G01N2446/80G01N2800/347
Inventor 王保君汤双双欧卫军牛青青陆燕子
Owner NANTONG EGENS BIOTECH
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