Detection kit of lipocalin related to neutrophil gelatinase

A technology for lipocalin and neutrophils, which is applied in biological testing, material inspection products, etc., can solve the problems of insufficient repeatability, large amount of specimens, and many influencing factors, and achieves improved detection sensitivity and detection range. Strong specificity and good stability

Inactive Publication Date: 2016-06-22
ZHEJIANG ZOYUN BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the manual operation of the ELISA method, not only the detection steps are cumbersome, the sample turnover time is long, but also there are many influencing factors, and the repeatability is not good enough. It is replaced by an automated chemiluminescent detection system, such as the Architecti2000SR automatic immune analyzer from Abbott Laboratories in the United States to measure urine NGAL (chemiluminescent microparticle immunoassay), the sample volume needs to be 150 μL, and the detection cycle is about 28 minutes; EDTA anticoagulant plasma or whole blood (double-antibody sandwich immunofluorescence chromatography), the detection of NGAL can be completed in 15 minutes, and the detection range is 50-2000ng / mL
However, these detection systems can only detect urine or plasma alone, the amount of specimens is large, and the instrument itself and supporting reagents are extremely expensive, which limits its routine clinical application

Method used

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  • Detection kit of lipocalin related to neutrophil gelatinase
  • Detection kit of lipocalin related to neutrophil gelatinase
  • Detection kit of lipocalin related to neutrophil gelatinase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1: kit preparation 1.

[0029] Preparation of Reagent 1:

[0030] To 50mmol / L phosphate buffer (pH=7.4), add turbidity agent at a final concentration of 0.1% (w / v) and NaN at a final concentration of 0.02% (w / v) 3 , fully stirred and mixed to prepare reagent 1, and stored at 4°C.

[0031] Preparation of Reagent 2:

[0032] 1) Add 0.2 mL of 5 mg / mL EDC and 0.2 mL of 5 mg / mL NHS to 1 mg of latex particles composed of two particle sizes; the latex particles are commercial products, purchased from JSR Corporation, with a diameter of 80 nm and 250nm, mixed with a volume ratio of 1:4;

[0033] 2) Shaking reaction at room temperature (20-25°C) for 30 minutes;

[0034] 3) Rinse the suspended particles with 50mmol / L phosphate buffer (pH=7.4) to remove excess unreacted EDC and NHS to obtain an activated latex suspension;

[0035] 4) Dilute the anti-neutrophil gelatinase-associated lipocalin antibody to 1 mg / mL with 50 mmol / L phosphate buffer (pH=7.4);

[0036] 5)...

Embodiment 2

[0042] Example 2: kit preparation 2.

[0043] Preparation of Reagent 1:

[0044] To 20mmol / L Tris buffer, add turbidity agent with a final concentration of 0.1% (w / v) and NaN with a final concentration of 0.02% (w / v) 3 , fully stirred and mixed to prepare reagent 1, and stored at 4°C.

[0045] Preparation of Reagent 2:

[0046] 1) Add 0.2 mL of 5 mg / mL EDC and 0.2 mL of 5 mg / mL NHS to 1 mg of latex particles composed of two particle sizes; the latex particles are commercial products, purchased from JSR Corporation, with a diameter of 80 nm and 250nm, mixed with a volume ratio of 1:4;

[0047] 2) Shaking reaction at room temperature for 30 minutes;

[0048] 3) Rinse the suspended particles with 50mmol / L phosphate buffer (pH=7.4) to remove excess unreacted EDC and NHS to obtain an activated latex suspension;

[0049] 4) Dilute the anti-neutrophil gelatinase-associated lipocalin antibody to 1 mg / mL with 50 mmol / L phosphate buffer (pH=7.4);

[0050] 5) Add 10 μL of diluted a...

Embodiment 3

[0056] Example 3: kit preparation 3.

[0057] Preparation of Reagent 1:

[0058] To 20mmol / L Tris buffer, add turbidity agent at a final concentration of 1% (w / v) and NaN at a final concentration of 0.02% (w / v) 3 , fully stirred and mixed to prepare reagent 1, and stored at 4°C.

[0059] Preparation of Reagent 2:

[0060] 1) Add 0.1 mL of 5 mg / mL EDC and 0.1 mL of 5 mg / mL NHS to 1 mg of latex particles composed of two particle sizes; the latex particles are commercial products, purchased from JSR Corporation, with a diameter of 80 nm and 250nm, mixed at a volume ratio of 1:1;

[0061] 2) Shaking reaction at room temperature for 30 minutes;

[0062] 3) Rinse the suspended particles with 50mmol / L phosphate buffer (pH=7.4) to remove excess unreacted EDC and NHS to obtain an activated latex suspension;

[0063] 4) Dilute the anti-neutrophil gelatinase-associated lipocalin antibody to 1 mg / mL with 50 mmol / L phosphate buffer (pH=7.4);

[0064] 5) Add 50 μL of diluted antibody ...

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Abstract

The invention provides a detection kit for neutrophil gelatinase-associated lipocalin, which is characterized in that it includes reagent 1 and reagent 2; the reagent 1 includes: 10-100 mmol/L buffer, 0.1 -1% w/v of turbidity agent, 0.01-0.05% w/v of preservative; said reagent 2 includes: 0.1-0.5% w/v labeled with anti-human neutrophil gelatinase-associated lipocalin sensitizing latex particles, 0.1-5% w/v stabilizer, 10-100 mmol/L buffer, 0.01-0.05% w/v preservative. The latex particles include latex particles of two different particle sizes, and the latex particles of the two particle sizes are mixed in an appropriate ratio. The kit has high sensitivity, wide linear range and good precision, and is suitable for automatic biochemical analyzers.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to a detection kit for detecting neutrophil gelatinase-associated lipocalin. Background technique [0002] With the continuous improvement of people's living standards and the gradual changes in living and eating habits, the incidence of kidney disease is increasing year by year. Kidney disease is mostly secondary to diabetes, hypertension, immune damage and bacterial infection. Its etiology is complicated, the course of the disease is protracted, and it increases the risk of cardiovascular disease, which has become a global public health problem. Kidney disease can be divided into two main syndromes - acute kidney injury (acute kidney injury, AKI) and chronic kidney disease (chronickidney disease, CKD). AKI refers to a group of clinical syndromes of sudden (1-7d) and persistent (>24h) renal function decline, manifested as azotemia, water-electrolyte and acid-base balance dis...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 梅义武刘兴叶佳颖
Owner ZHEJIANG ZOYUN BIOTECH
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