Kit and method for detecting hepatitis B virus surface antigen (HBsAg)

A technology for hepatitis B virus and detection reagents, which is applied to measurement devices, instruments, scientific instruments, etc., can solve the problems of detecting HBsAg without metalloporphyrin complexes, and achieves overcoming high non-specific background signal and low labeling rate, The effect of reducing systematic errors and improving detection sensitivity

Inactive Publication Date: 2011-09-14
HANGZHOU NORMAL UNIVERSITY
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  • Kit and method for detecting hepatitis B virus surface antigen (HBsAg)
  • Kit and method for detecting hepatitis B virus surface antigen (HBsAg)

Examples

Experimental program
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Embodiment 1

[0027] Embodiment 1 Novel water-soluble A with high labeling rate 3 Preparation of B-type metalloporphyrin-labeled antibody

Embodiment 2A3

[0028] Add 10 mg of N-hydroxysuccinimide (NHS) into 1 mL of aqueous solution containing 5 mg of metalloporphyrin, stir magnetically at 20°C for 10 min, then add 4 mL of aqueous solution containing 2.0 mg of hepatitis B virus surface antibody (HBsAb), and continue at 20°C Stir for 3 hours, the product was centrifuged at 3000r / min at 4°C for 20 minutes, the supernatant was salted out with 10 g of solid ammonium sulfate, and the precipitate after salting was dialyzed with phosphate buffer (pH 7.0) for 1 hour until the permeate was free. Color and freeze-dry to separate metalloporphyrin-labeled HBsAb with a high labeling rate, which is the detection reagent for HBsAg. Example 2A 3 Chemiluminescence immunodetection of HBsAg labeled with type B metalloporphyrin HBsAb

[0029] (1) Coating: Use pH 7.0, 0.1mol / L phosphate buffer solution to dilute 125 μL 1.0 mg / mL HBsAb aqueous solution to 50.0 mL, prepare 2.5 μg / mL coating solution, add 100 μL per well to 96-well white Place in a cl...

Embodiment 3

[0033] Example 3A 3 Chemiluminescence immunodetection of HBsAg labeled with type B metalloporphyrin HBsAb

[0034] (1) Coating: Dilute 50 μL of 1.0 mg / mL HBsAb aqueous solution to a volume of 50.0 mL with pH 7.4, 0.1 mol / L phosphate buffer solution to prepare 1.0 μg / mL coating solution, and add 200 μL to each well of 96-well white In a bottom-transparent polystyrene microwell plate, place at 37°C for 60 minutes, discard the coating solution, add 200 μL of the above-mentioned phosphate buffer solution containing 5% bovine serum albumin (BSA) to each well as a blocking solution, block at 37°C for 1 hour, Wash with pH 7.4, 0.1mol / L phosphate buffer solution, soak for 3 minutes each time, wash the plate 3 times, vacuum dry at 30°C and put it in a sealed bag for vacuum storage to prepare a microwell plate coated with HBsAb;

[0035] (2) Adding samples: divide the microwell plate coated with HBsAb prepared in step (1) into a standard group and a sample group to be tested, and the c...

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Abstract

The invention discloses a kit and a method for detecting hepatitis B virus surface antigen (HBsAg). The detection kit is used for marking hepatitis B virus surface antibody (HBsAb) by using metalloporphyrin obtained by using water-soluble A3B type metalloporphyrin to mark the HBsAb. The detection kit and the detection method have the advantages that: (1) free metalloporphyrin molecules are separated from a coupled substance easily after an A3B type metalloporphyrin complex is coupled with the HBsAb to obtain a high-purity labelled antibody, so that non-specific adsorption background signals are reduced when the HBsAg is detected by chemical luminescence immunodetection, and detection sensitivity is greatly improved; (2) the method is simple, convenient and quick; automation is realized easily; and specificity of immunoreaction, high sensitivity of chemical luminescence reaction and stability of a chemical marker are achieved; and (3) a new method is provided for detecting the HBsAg quickly and accurately, and preventing and treating the hepatitis B virus.

Description

(1) Technical field [0001] The invention relates to a detection reagent for hepatitis B surface antigen HBsAg and a method for detecting HBsAg, in particular to a water-soluble A 3 Type B metalloporphyrin-labeled hepatitis B surface antibody HBsAb is used as a detection reagent for HBsAg and a method for detecting HBsAg. (2) Background technology [0002] Hepatitis B (HBV) virus infection is a global problem and a serious threat to human health. HBV is an important pathogenic factor causing acute and chronic hepatitis, liver cirrhosis and primary hepatocellular carcinoma. Hepatitis B virus surface antigen (HBsAg) has been the most important marker in the detection of hepatitis B virus infection, and is one of the early diagnostic indicators of hepatitis B. At present, the most commonly used HBsAg determination methods in China are colloidal gold immunochromatography (GICA) and enzyme-linked immunosorbent assay (ELISA): the former has the advantages of quickness and convenie...

Claims

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Application Information

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IPC IPC(8): G01N33/576C07D487/22
Inventor 陈灿玉黄志坚袁通张现侠何睿李霞梁媛媛焦艳华
Owner HANGZHOU NORMAL UNIVERSITY
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