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Immunoturbidimetric NGAL detection reagent and method

An immunoturbidimetric and detection reagent technology, which is applied in the field of medical immunodiagnosis, can solve the problems of unfavorable promotion and implementation in small and medium-sized hospitals, reduce test precision, time-consuming and laborious, etc., and achieve shortened test time, simplified operation, and good applicability of reagents Effect

Inactive Publication Date: 2018-05-29
SUZHOU KANGHESHUN MEDICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The operation of diluting and mixing must be automatically operated by the machine on the fully automatic biochemical analyzer (the common instrument platform of the third-level hospital), but on the small semi-automatic platform, especially the portable platform of POCT (hospitals below the third level and the medical treatment at the county and township level) Commonly used by service units) tests need to be performed manually by operators, which brings additional operation steps, introduces more operational errors, and reduces the precision of the test, which is not conducive to the promotion and implementation of the test in small and medium-sized hospitals
Another disadvantage of the NGAL latex-enhanced immune turbidimetric reagents on the market is that it can only measure the NGAL content in serum or plasma samples, and the whole blood samples must be pre-treated to extract serum, which is time-consuming and laborious, and has poor repeatability. Very inconvenient in clinical practice

Method used

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  • Immunoturbidimetric NGAL detection reagent and method
  • Immunoturbidimetric NGAL detection reagent and method
  • Immunoturbidimetric NGAL detection reagent and method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] 1) Preparation of latex reagent

[0043] The surface carboxylated microspheres with a particle size of 100nm were diluted to 1% in MES (25mM, pH=6.5) buffer solution and stirred at room temperature. Put in EDC solid powder, keep the molar ratio of EDC and carboxyl group at 10:1, stir for 2 hours and then centrifuge at 10000RPM for 10 minutes. The latex was washed twice with MES (25mM, pH=6.5) buffer solution and resuspended. Add NGAL antibody, the ratio of antibody mass to latex mass is 0.025:1, and stir for 2 hours. Then centrifuge at 10,000 RPM for 15 minutes, and discard the supernatant. Latex was resuspended in storage solution (50mM Gly pH=7.4 + 0.5%BSA + 0.1%ProClin 300 + 0.5% Triton X-100 + 1% trehalose + 2.7% sodium chloride) and ultrasonically dispersed, diluted to a latex concentration of 0.2 % or so spare.

[0044] 2) Preparation of standard products

[0045] The commercially available NGAL antigen standard was prepared into a series of standard products...

Embodiment 2

[0051] 1) Preparation of latex reagent

[0052] The surface carboxylated microspheres with a particle size of 100nm were diluted to 1% in MES (25mM, pH=6.5) buffer solution and stirred at room temperature. Put in EDC solid powder, keep the molar ratio of EDC and carboxyl group at 10:1, stir for 2 hours and then centrifuge at 10000RPM for 10 minutes. The latex was washed twice with MES (25mM, pH=6.5) buffer solution and resuspended. Add NGAL antibody, the ratio of antibody mass to latex mass is 0.025:1, and stir for 2 hours. Then centrifuge at 10,000 RPM for 15 minutes, and discard the supernatant. Latex was resuspended in storage solution (50mM Gly pH=7.4 + 0.5%BSA + 0.1%ProClin 300 + 0.5% Triton X-100 + 1% trehalose + 2.7% sodium chloride) and ultrasonically dispersed, diluted to a latex concentration of 0.05 % or so spare.

[0053] 2) Preparation of standard products

[0054] The commercially available NGAL antigen standard was prepared into a series of standard product...

Embodiment 3

[0060] Analytical sensitivity of reagents (blank limit)

[0061] Using the reagents in Example 1, select the zero-value standard as a blank sample test, and test the absorbance value on an automatic biochemical analyzer (Mindray BS-400) at a temperature of 37°C, a wavelength of 546nm, and an optical path of 1cm. The test was repeated 20 times, and the mean (X) and standard deviation (SD) of the 20 test results were calculated according to the standard curve established in Example 1, and X+2SD was calculated as the analytical sensitivity of the reagent. The test results are shown in Table 1, showing that the analytical sensitivity of the reagent is 8.456 ng / mL. The clinical reference value for detection of neutrophil gelatinase-associated lipocalin (NGAL) is about 200 ng / mL, and the sensitivity of the reagent of the present invention is an order of magnitude smaller than the reference value, which fully meets the needs of use.

[0062] Example 1

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Abstract

The invention discloses a latex-enhanced immunoturbidimetric NGAL (neutrophil gelatinase associated lipocalin) detection reagent. The latex-enhanced immunoturbidimetric NGAL detection reagent is a single reagent, uses NGAL antibody-labeled latex particles as a main component, and further comprises a buffer solution, a surfactant, salt, a stabilizer, a suspending agent and a preservative. The invention further discloses a method for detecting the concentration of NGAL in a blood sample by virtue of a transmitting or scattering turbidimetric principle by using the latex-enhanced immunoturbidimetric NGAL detection reagent. The latex-enhanced immunoturbidimetric NGAL detection reagent adopts the single reagent, is easy to operate, free of need of mixing, high in sensitivity and wide in linearrange, can directly measure various samples such as whole blood, serum and plasma, and can be widely applied to various transmitting or scattering analyzers, including an ordinary biochemical analyzer, a specific protein analyzer and the like.

Description

technical field [0001] The invention belongs to the field of medical immunodiagnosis, and in particular relates to the preparation and application of a latex-enhanced immune turbidimetric detection reagent for neutrophil gelatinase-associated lipocalin (NGAL). Background technique [0002] Neutrophil gelatinase-associated lipocalin (NGAL), also known as human lipocalin 2 (lipocalin2, Ln2) or siderocalin, is a member of the human lipocalin family new member. In recent years, NGAL has attracted much attention as a new marker of renal injury. NGAL in serum and urine will increase significantly in early acute kidney injury, chronic kidney disease, cardiorenal syndrome and other diseases, and its level monitoring has good diagnostic value in judging the severity, curative effect and prognosis of the disease. Acute kidney injury (AKI) is a clinical syndrome caused by a sudden decline in renal function in a short period of time (hours to days) caused by various reasons. It is a c...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/546
CPCG01N33/6893G01N33/54313G01N2333/47G01N2800/347
Inventor 陈胜胜刘向晖王明
Owner SUZHOU KANGHESHUN MEDICAL TECH
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