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Kit for detecting anti-GP73 antibody in serum

A kit and serum technology, applied in the field of medicine and biology, can solve the problems of unreported diagnosis of liver cancer, etc., and achieve the effect of kit improvement, high medical value, and good specificity

Inactive Publication Date: 2014-12-17
GUANGXI MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Although there have been reports on the detection of GP73 autoprotein in the serum of liver cancer patients, when the liver cancer tissue releases the tumor-associated marker GP73 protein into the blood, how does the body produce an immune response to the protein and how does the body produce antibodies against the protein? Its role in the diagnosis of liver cancer has not been reported

Method used

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  • Kit for detecting anti-GP73 antibody in serum
  • Kit for detecting anti-GP73 antibody in serum
  • Kit for detecting anti-GP73 antibody in serum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Embodiment 1 detects the preparation method of the kit of anti-GP73 antibody in serum

[0065] 1. Production of GP73 protein antigen

[0066] The GP73 protein antigen in the kit of the present invention is artificially synthesized, and the amino acid sequence list of the GP73 protein is shown in SEQ ID NO:1. The preparation process of GP73 protein antigen among the present invention is as follows:

[0067] (1) Design primers according to the gene of GP73 protein antigen, including upstream primer SEQID NO: 3'-GGAACGGTACCCACCATCATCATCATCAGGCTGCCCTGTCAGTGAGCCAGGAAAA-3', downstream primer SEQID NO: 4: 5'-GGAACGTCGACTCAGAGTGTATGATTCCGCTTTTCACGCTGATCA

[0068] AGTAAATT-3', use RT-PCR to amplify the GP73 gene, the specific steps are: take a sterile PCR tube, add 2×TaqMasterMix 25μl in sequence, each 2μl of upstream primer and downstream primer, 1μl of reverse transcription product of HepG2 cell total RNA, ddH 2 Make up to 50 μl with O; gently centrifuge and mix the added PC...

Embodiment 2

[0089] Example 2 Detection of Anti-GP73 Antibody Kit in Serum Using Method

[0090] The present invention also provides a detection method for detecting anti-GP73 antibody content in serum using the above kit, the method comprising the following steps:

[0091] (1) Dilute the rabbit anti-human GP73 polyclonal antibody standard (320ng / ml) with 0.01mol / l PBS diluent to 160ng / mL, 80ng / mL, 40ng / mL, 20ng / mL, 10ng / mL five concentration.

[0092] (2) Create a standard curve for the relationship between absorbance and antibody concentration:

[0093] ① Coating: GP73 protein antigen was diluted to 4 ng / ml with coating buffer 0.5% bovine serum albumin (BSA), and coated overnight at 4°C.

[0094] ②Sealing: wash the plate with washing solution, pat dry, and block with 0.5% BSA for 1h.

[0095] ③ Add standard protein: wash the plate with washing solution, pat dry, add 50 μl of rabbit anti-human GP73 polyclonal antibody standard with different dilution concentrations above to the microwe...

Embodiment 3

[0102] The detection result of embodiment 3 kit to normal person specimen, hepatitis, liver cirrhosis specimen, and liver cancer specimen

[0103] The inventor detected GP73 antibody in serum of 100 routine normal control groups, 100 routine hepatitis and liver cirrhosis groups (i.e. benign liver disease group), and 127 routine primary liver cancer groups with the kit, and the results showed that the antibody concentrations of these three groups were 46.36 ± 13.18ng / ml, 140.34±18.38ng / ml, 259.23±86.72ng / ml, such as image 3 shown. Serum GP73 antibody concentration in patients with primary liver cancer was significantly higher than that in the normal group and hepatitis and cirrhosis group (P<0.05), and there was also a significant statistical difference between the serum GP73 antibody concentration in patients with hepatitis and cirrhosis and the normal group (P<0.05 ).

[0104] Taking the normal group as the liver cancer control group, the abnormal serum cutoff value (cutof...

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Abstract

The invention discloses a kit for detecting an anti-GP73 antibody in serum, and belongs to the technical fields of medicines and biology. The kit comprises GP73 protein antigen, confining liquid, GP73 protein antigen of an enzyme label, a rabbit anti-human GP73 polyclonal antibody standard, a coating buffer, a developing solution, a scrubbing solution, a stop solution and a diluent, wherein the GP73 protein antigen has an amino acid sequence as shown in a sequence table SEQIDNO:1. According to the kit, the concentration of the anti-GP73 antibody in the serum of a patient can be accurately calculated, the existential level of the anti-GP73 antibody in the serum is really reflected, the kit can be applied to diagnosis of primary liver cancer in clinic, and the process of molecular change of the tumor can also be reflected by detecting the level of the anti-GP73 antibody, thereby judging the autoimmune response capacity of a silk body. The kit disclosed by the invention also has the advantages of good specificity, and high sensitivity, accuracy and precision.

Description

technical field [0001] The invention relates to the field of medical biotechnology, in particular to a kit for detecting anti-GP73 antibodies in serum, and a detection method for using the kit to detect serum marker anti-GP73 antibodies in patients with liver cancer. Background technique [0002] The incidence of liver cancer ranks fourth in the world and the second in China, and it is increasing year by year worldwide, with a total incidence of more than 560,000. Liver cancer is relatively hidden at the beginning of its onset, and it is difficult to detect it clinically. The detection of clinical diagnosis is generally late, and the treatment effect is poor. Early detection of liver cancer and timely treatment can improve the 5-year survival rate of patients. Therefore, early detection of liver cancer has important clinical significance significance. [0003] Currently, liver cancer detection methods include liver ultrasound image analysis and serological marker detection,...

Claims

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Application Information

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IPC IPC(8): G01N33/574G01N33/535
CPCG01N33/535G01N33/57438
Inventor 周素芳薛冰滢
Owner GUANGXI MEDICAL UNIVERSITY
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