Detection kit for human serum amyloid A protein

A technology of amyloid and detection kits, applied in the field of medical immunology, can solve the problems of increasing the amount of antibodies and production costs, loss of antibody binding force, false positive and false negative test results, etc., to improve accuracy and credibility , reduce the amount of antibody used, and reduce production costs

Inactive Publication Date: 2016-03-02
ZHEJIANG ZOYUN BIOTECH
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AI Technical Summary

Problems solved by technology

[0004] The existing SAA detection method is mainly enzyme-linked immunoassay. Due to the long cycle of enzyme-linked immunoassay, it cannot be used on automatic biochemical analyzers, and it is not suitable for rapid detection in emergency. The turbidimetric method is simple and economical, and can be used on the automatic biochemical analyzers in most hospitals, especially in emergency departments, where rapid quantitative detection can be achieved. After the reaction, aggregated particles are formed, and at a certain wavelength, the content of the analyte in the sample can be measured by measuring the turbidity generated by the aggregate
[0005] However, when the serum amyloid A antibody is coated on the latex, physical adsorption and chemical coupling methods can generally be used. The stability of the sensitized latex particles obtained by the physical adsorption method is worse than that of the chemical coupling method, and the ...

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  • Detection kit for human serum amyloid A protein

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Embodiment 1 Preparation of Human Serum Amyloid A Detection Kit

[0027] The kit of the present invention relates to the main raw materials of reagents as follows:

[0028] 1. Serum amyloid A antibody: the titer measured by indirect ELISA method is 1:100000.

[0029] 2. Latex: The present invention only exemplarily uses polystyrene latex particles with a diameter of 100-200 nm and carboxyl groups to carry out experiments.

[0030] The preparation of the main reagent of this embodiment is as follows:

[0031] Reagent R1: ammonium chloride buffer containing 1.5% PEG6000 (polyethylene glycol 6000), 0.8% NaCl. The reagent is a colorless transparent solution.

[0032] Reagent R2: latex particles were sensitized with anti-serum amyloid A antibody. The reagent is a milky white solution.

[0033] Specific steps are as follows:

[0034] 1. Take 1ml (100mg / ml) latex, wash 3 times with 0.1M (mol / L), pH5.0 MES solution (2-morpholineethanesulfonic acid buffer), and ultrasonica...

Embodiment 2

[0043] Embodiment 2 Determination of human serum amyloid A

[0044] Detection tool: Hitachi 7080 automatic analyzer.

[0045] Analysis method: two-point endpoint method; main wavelength: 546nm, secondary wavelength: 700nm: sample volume: 2.0ul; R1: 240ul; R2: 60ul; calibration method: Logit-log5p; reaction direction: rising; measurement temperature: 37°C; Add R2 when the sample is mixed with R1 for 5 minutes, read the absorbance A1 at the first point immediately, and read the absorbance A2 after 5 minutes. The reaction absorbance was calculated as the calibrated difference between A2 and A1.

[0046] Calculation method: multi-point calibration, the dose / response curve is made according to the absorbance and reference serum value, and the sample content can be calculated on the dose / response curve according to its absorbance value.

Embodiment 3

[0047] Example 3 Performance Evaluation of Human Serum Amyloid A Detection Kit

[0048] 1. Analytical Sensitivity Assessment

[0049] Use a 200mg / L sample to measure the kit, repeat the measurement 3 times, and convert the change in absorbance at the concentration of 198.9mg / L specified in the kit. The results show that the absorbance at the concentration of 198.9mg / L is 0.0730 (see Table 1).

[0050] Table 1

[0051] Measurement times Determination of the mean absorbance Absorbance at 198.9mg / L concentration 3 0.0734 0.0730

[0052] 2. High value linear evaluation

[0053] Mix the samples with SAA concentrations of 0mg / L and 400mg / L in different proportions, and repeat the measurement 3 times for each concentration (see Table 2)

[0054] The highest detection range of the detection kit of the present invention can reach 400mg / L, and the judgment basis is r2≥0.990.

[0055] Table 2

[0056] sample ratio Theoretical valuemg / L 3 times de...

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Abstract

The invention discloses a detection kit for human serum amyloid A protein. The detection kit is characterized by comprising a reagent R1 and a reagent R2, the reagent R1 is an appropriate buffer solution, and the reagent R2 is prepared by putting polystyrene latex particles sensitized by a human serum amyloid A protein antibody into the appropriate buffer solution. The detection kit has the advantages of being simple and fast in detection, high in sensitivity, good in accuracy, high in anti-jamming capability and low in production cost.

Description

technical field [0001] The invention belongs to the field of medical immunology, and relates to an immunodetection reagent, and further, the invention relates to a human serum amyloid A detection kit. Background technique [0002] Serum amyloid A (serumamyloidAprotein, SAA) is an acute phase response protein, which belongs to the heterogeneous protein in the apolipoprotein family, with a relative molecular weight of about 12,000. In the acute phase response, stimulated by IL-1, IL-6 and TNF, SAA is synthesized in the liver by activated macrophages and fibroblasts, which can increase to 100-1000 times the initial concentration, but the half-life Short, only about 50 minutes. Serum amyloid A is associated with high-density lipoprotein (HDL), which regulates HDL metabolism during inflammation. A particularly important property of serum amyloid A is that its degradation products can be deposited in different organs as amyloid A (AA) fibrils, which is a serious complication in ...

Claims

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Application Information

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IPC IPC(8): G01N33/92G01N33/546
CPCG01N33/54313G01N33/92G01N2333/775
Inventor 梅义武刘兴李静
Owner ZHEJIANG ZOYUN BIOTECH
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