PSMD4 protein ELISA detection kit as well as detection method and application thereof

A detection kit, PSMD4 technology, applied in the fields of immunology and biology, can solve problems such as simplicity

Active Publication Date: 2015-12-30
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there is no literature report on the accurate detection of PSMD4 protein in liver cancer serum samples; there is no accurate, simple and sensitive detection method for PSMD4 protein content in liver cancer serum samples; there is no literature report on PSMD4 protein ELISA Detection kit and application of such kit in serological diagnosis of liver cancer

Method used

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  • PSMD4 protein ELISA detection kit as well as detection method and application thereof
  • PSMD4 protein ELISA detection kit as well as detection method and application thereof
  • PSMD4 protein ELISA detection kit as well as detection method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Preparation of detection ELISA plate: Coating buffer 1×PBS, pH: 7.4 Dilute the antibody (rabbit anti-human PSMD4 polyclonal antibody) to 2.0ug / ml, add 100 μl to each well of the ELISA plate, Seal the plate with blocking solution containing 3% bovine serum albumin + PBS solution and incubate overnight at 4°C for later use.

[0075] Detection of serum samples and recombinant proteins: serum samples were diluted 1:1 with sample diluent 1×PBS (pH:7.4), added at a volume of 100 μl / well, and incubated at room temperature for 2 hours; PSMD4 recombinant protein was diluted with sample diluent In different concentration gradients, add samples at a volume of 100 μl / well, and incubate at room temperature for 2 hours; shake off the liquid in each well, and add ELISA plate washing solution (1×PBS solution containing 0.05% Tween-20), each well 300 μl, washed 3-5 times, and dried; Dilute the PSMD4 detection antibody to 0.5 μg / ml, 100 μl per well, incubate at room temperature for 1 hou...

Embodiment 2

[0076] Example 2: Optimization

[0077] 1. Optimization of coating antibody concentration:

[0078] Select different coating antibodies rabbit anti-human PSMD4 polyclonal antibody (PTG company and Abnova company), choose different concentrations (7.5μg / ml, 5μg / ml, 2.0μg / ml, 1μg / ml, 0.5μg / ml, 0.25 μg / ml) coated ELISA plate, detected according to the operating steps in Example 1, adding known PSMD4 protein standards of different concentrations. According to the obtained OD value, select the blank group with the smallest OD value and the closest linear relationship between the OD value and the standard protein concentration as the best antibody (PTG company). The coating antibody concentration is 2.0 μg / ml.

[0079] 2. Optimization of blocking solution:

[0080] Choose different blocking solutions, containing 1% bovine serum albumin + PBS solution, containing 2% bovine serum albumin + PBS solution, containing 3% bovine serum albumin + PBS solution, containing 4% bovine serum al...

Embodiment 3

[0087] Embodiment 3: Specificity and sensitivity evaluation of the ELISA kit of PSMD4 protein

[0088] 1. Specificity test

[0089] The ELISA method established in Example 1 was used to detect the serum samples of 100 liver cancer patients, 50 normal people and 20 patients with liver cirrhosis (the healthy people were selected from the physical examination samples of our hospital staff; The pathological diagnosis was primary liver cancer; the liver cirrhosis sample was from the Department of Infectious Diseases of Changhai Hospital, and the clinical diagnosis was liver cirrhosis), PSMD4 recombinant protein was used as a positive control, and a standard curve was drawn according to the obtained standard OD value and the corresponding concentration to calculate the Serological levels of PSMD4 protein in .

[0090] The result is as figure 2 As shown, the serum level of PSMD4 protein in liver cancer group (Median388.2pg / mL, IQR348.3pg / ml) was significantly higher than that of l...

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Abstract

The invention belongs to the fields of immunology and biotechnology, and particularly relates to a PSMD4 protein ELISA detection kit as well as a detection method and application to serological diagnosis for liver cancer of the kit. According to extensive and deep research of the inventor, the expression level of PSMD4 protein in the serum of liver cancer patients can be detected according to an enzyme-linked immunosorbent assay (ELISA) method, and the PSMD4 protein is proved for the first time in the serum of the liver cancer patients. The kit comprises an ELISA plate coated with a PSMD4 antibody, a PSMD4 antigen detection antibody, an ELIAS secondary antibody, standard protein and the like. The invention further provides the detection method and application of the kit. The kit is simple and convenient to operate, can detect the content of the PSMD4 protein in the serum of a liver cancer patient correctly and highly sensitively, and provides a novel method for clinical examination and basic research.

Description

technical field [0001] The invention belongs to the field of immunology and biotechnology, and in particular relates to an ELISA detection kit for PSMD4 protein, a detection method thereof and an application in serological diagnosis of liver cancer. Background technique [0002] Primary hepatocellular carcinoma (hepatocellular carcinoma, HCC) is one of the most common malignant tumors in my country, and the annual new cases account for about 50% of the world. Since liver cancer is mostly in the middle and late stage when symptoms appear, the recurrence and metastasis rate after resection is high. Therefore, early diagnosis of liver cancer is of great significance to prolong the survival time of patients and reduce the mortality of liver cancer. [0003] Currently, the diagnosis of HCC mainly relies on laboratory tests, imaging tests, and histological examinations of liver biopsy. Imaging diagnosis plays an important role in the diagnosis of liver cancer, but it has certain...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/573G01N33/577G01N33/543
CPCG01N33/543G01N33/573G01N33/57438G01N33/57484G01N2333/948G01N2800/085
Inventor 王红阳丁劲刘娜李晓峰
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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