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Kit for detecting SAA (serum amyloid A) with spatial proximity chemiluminescence method and detection method of kit

A detection kit, chemiluminescence technology, used in chemiluminescence/bioluminescence, biological testing, analysis by chemical reaction of materials, etc., can solve the problem of unstable detection results, many influencing factors, poor repeatability, etc. problem, to achieve the effect of eliminating luminous interference, simple production process, and easy to fully automate

Inactive Publication Date: 2019-01-04
无锡壹闪生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Its operation is complicated and there are many influencing factors, resulting in unstable test results and poor repeatability

Method used

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Examples

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Effect test

preparation example Construction

[0027] 1. Preparation of calibrator

[0028] (1) Preparation of calibration product dilution: Weigh 14.1 g of potassium dihydrogen phosphate, sodium dihydrogen phosphate (2H 2 O) 3.0g, add ultrapure water to dissolve, Proclin-300 0.5 ~ 1mL, after mixing, add ultrapure water to make up to 1000mL, obtain the calibration product dilution, store at 2 ~ 8 ℃ for later use.

[0029] (2) Preparation of calibrator: the concentration of calibrator is 0, 15ng / mL, 45ng / mL, 135ng / mL, 400ng / mL, 800ng / mL, dilute the purified serum amyloid A to the corresponding concentration with calibrator diluent , Store at 2-8°C for later use.

[0030] 2. Preparation of enzyme markers

[0031] (1) Weigh 5 mg of HRP and dissolve it in 1 mL of distilled water, add 0.2 mL of freshly prepared 0.1M sodium periodate solution to the supernatant, stir at room temperature in the dark for 20 min, put the above solution into a dialysis bag, and adjust to 1 mM pH 4.4 Dialyzed against sodium acetate buffer overnight ...

Embodiment 1

[0049] This embodiment provides a detection kit for serum amyloid A, including: enzyme markers, luminescent markers, auxiliary agents, triggers, and calibrators; wherein, the calibrator includes no less than 6 different concentrations of SAA antigens Calibrator and 0.1M phosphate buffer; enzyme markers include peroxidase-labeled SAA detection antibody and 0.05M phosphate buffer; luminescent markers include 9,10-dihydroacridine-labeled SAA capture antibody and 0.05 M Tris buffer; the components of the auxiliary agent include luminescent auxiliary agent and pH 6.0 citrate buffer; the trigger agent is selected from 0.05M pH 8.0 Tris-HCl buffer.

[0050] 1. Preparation of calibrator

[0051] (1) Preparation of calibration product dilution: Weigh 14.1 g of potassium dihydrogen phosphate, sodium dihydrogen phosphate (2H 2 O) 3.0g, add ultrapure water to dissolve, Proclin-300 0.8mL, after mixing, add ultrapure water to make up to 1000mL to obtain the calibration product dilution, st...

Embodiment 2

[0066] This embodiment provides a method for detecting serum amyloid A using a serum amyloid A detection kit in a spatially adjacent chemiluminescence method, including steps:

[0067] S1: Add 25 μL of standard substance, 25 μL of peroxidase-labeled SAA detection antibody and 25 μL of luminescent marker to the chemiluminescence reaction tube respectively.

[0068] S2: React in a 37°C incubator for 15 minutes.

[0069] S3: Add 5 μL of auxiliary agent to the chemiluminescent reaction tube, shake and mix, and let it stand for 1-2 minutes; then add 75 μL of trigger agent, shake and mix, detect immediately, and read the signal value.

[0070] S4: Carry out logistic four-parameter fitting on the concentration and luminescence value of the calibrator, and calculate the sample concentration through the luminescence value of the sample.

[0071] The invention adopts the spatial proximity chemiluminescence method to detect the serum amyloid A, and the sensitivity can reach 1mg / L. SAA ...

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Abstract

The invention relates to a kit for detecting SAA (serum amyloid A) with a spatial proximity chemiluminescence method and a detection method of the kit. The kit comprises an enzyme marker, a luminescent marker, an adjuvant, a trigger and a calibrator, wherein the calibrator contains calibration products of SAA antigens with different concentrations and a 0.1 M phosphate diluent; the enzyme marker contains components of a peroxidase labeled SAA detection antibody and a 0.05 M phosphate buffer; the luminescent marker contains components of a 9,10-dihydracridine labeled PCT capture antibody and 0.05 M Tris buffer; the adjuvant contains components of a luminescent adjuvant and a citrate buffer. A reagent spatial proximity luminescence analysis detection technology is adopted serves as a true homogeneous chemiluminescence technology, cleaning and a carrier are not needed, a coating process is omitted, and detection sensitivity and specificity are high, so that detection results are more realand reliable; meanwhile, reaction time and reaction steps are optimized, and operation is simpler.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a kit for detecting serum amyloid A by space proximity chemiluminescence method and a detection method thereof. Background technique [0002] Infectious diseases are common clinical diseases and frequently-occurring diseases, which are mainly inflammatory reactions caused by pathogens such as bacteria, viruses and / or fungi invading the body, and most of them are accompanied by fever symptoms. While antibacterial drugs have cured and saved the lives of many patients, their irrational use and abuse have also led to many adverse consequences, such as the emergence of extensive and serious drug resistance in bacteria. [0003] Serum amyloid is a polypeptide composed of 104 amino acids. Its relative molecular mass in the natural state is about 12000-14000. The gene of serum amyloid is located on chromosome 11. It is a sensitive inflammatory marker discovered in recent years. It is a An a...

Claims

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Application Information

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IPC IPC(8): G01N21/76G01N33/68G01N33/577G01N33/535
CPCG01N21/76G01N33/535G01N33/577
Inventor 奚伟红史兵伟廖鸳鸯朱丹丹
Owner 无锡壹闪生物科技有限公司
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