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Application of TIGAR protein in plasma as molecular marker in preparation of kit for predicting and diagnosing liver cancer recurrence

A molecular marker and kit technology, applied in the field of biotechnology detection, can solve the problems of increasing patient time, increasing the number of waiting lists, increasing the risk of liver transplantation dissemination and recurrence, etc.

Pending Publication Date: 2022-03-11
THE FIRST AFFILIATED HOSPITAL OF XIAMEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, a fatal "pain point" of liver transplantation is the shortage of donors, which has produced a series of chain reactions, first leading to a continuous increase in the number of people on the waiting list, thus increasing the time for patients from deciding to transplant to receiving the liver transplant itself; Secondly, during the waiting period, liver cancer cells will not wait obediently, but will continue to develop, eventually causing many liver cancer patients waiting for transplantation to withdraw from the list. This probability is proportional to time; finally, tumors occur during the waiting process Progression, but not enough to delist, but the condition in any case greatly increases the risk of dissemination and recurrence after liver transplantation

Method used

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  • Application of TIGAR protein in plasma as molecular marker in preparation of kit for predicting and diagnosing liver cancer recurrence
  • Application of TIGAR protein in plasma as molecular marker in preparation of kit for predicting and diagnosing liver cancer recurrence
  • Application of TIGAR protein in plasma as molecular marker in preparation of kit for predicting and diagnosing liver cancer recurrence

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0039] Plasma ELISA assay. Methods as below:

[0040] (1) Plasma thawing: The fresh frozen plasma of the liver cancer group and the control group, which were stored at -80°C, were taken out, and placed in a 4°C ice box to thaw.

[0041] (2) Use human fructose-2,6-bisphosphatase TIGAR ELISA kit (Wuhan Yiaibo Technology Co., Ltd., number: E5848h), the sensitivity of the kit is 0.085ng / mL, and the detection range is 0.15 -10.0 ng / mL. Take the kit out of the -20°C refrigerator and allow the reagents inside to equilibrate to room temperature.

[0042] (3) Gradiently dilute the standard product: first add the lyophilized standard product to the sample diluent to dilute 1.0mL, dissolve and mix well, and then let it stand for 10min. At this time, the concentration of the solution is 10.0ng / mL. 5.0ng / mL, 2.5ng / mL, 1.25ng / mL, 0.62ng / mL, 0.31ng / mL and 0.15ng / mL, put them into marked EP tubes respectively, and use the sample diluent as 0ng / mL , and its stock solution is used as the hi...

Embodiment 2

[0064] A. Extraction of total protein

[0065] Tissue homogenate: take 100mg of cancer tissue and paracancerous tissue respectively from fresh frozen tissue, wash away the blood water with 1×PBS, blot the water with filter paper; cut the tissue on ice with sterile scissors, add to a 1.5ml centrifuge tube Put the prepared tissue lysate into the shredded tissue. Homogenized steel balls were soaked with 75% alcohol for 30 minutes, washed with pure water for 3 times, each time for 3 minutes; ddH 2 O soak for later use. Wipe the steel balls with filter paper and add them to the centrifuge tube containing tissue lysate and tissue. Turn on the homogenizer 20 minutes in advance. After the internal temperature of the homogenizer drops below 4°C, put it into the prepared centrifuge tube and set up the frozen homogenizer. The parameters are 45Hz, 60s, 5 cycles, and stand still for 10min after the end. Take the centrifuge tube out of the homogenizer and put it into the low-temperature ...

Embodiment 3

[0105] The fixed and dehydrated tissue samples were divided into two parts. One part was processed according to the procedures of transparency, wax dipping and embedding. The parameters of the paraffin microtome were set at 4 μm for continuous sectioning. Adhesive glass slides were used to paste the tissues, and the cut paraffin sections were placed for 4 Store at ℃; another copy of OCT was embedded, and the parameters of the cryostat were set at 4 μm for continuous sectioning, and the slides were pasted to paste the tissue, and the cut frozen sections were stored in a -80°C refrigerator.

[0106] A. Hematoxylin-eosin staining (HE staining)

[0107] 1. Paraffin sections are baked: set the temperature at 60°C for 2-2.5 hours. 2. After the paraffin is melted, carry out the dewaxing hydration step. First, prepare 2 parts of xylene, 2 parts of absolute ethanol, 1 part of 95%, 80%, 75% and 50% ethanol, 1 part of distilled water, and each of the above liquids 250-350ml (subject to ...

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Abstract

The invention discloses application of TIGAR protein in plasma as a molecular marker in preparation of a kit for predicting and diagnosing liver cancer recurrence. The research finds that the content of the TIGAR protein in the blood plasma is obviously increased in cancer patients, the cutoff value for diagnosing liver cancer recurrence is 0.42 ng / ml, the area under the curve (ROC-AUC) is 0.802, the sensitivity is 100%, and the specificity is 60.47%; the ROC-AUC of the combined diagnosis of the recurrence with the AFP is 0.826, which is higher than that of the TIGAR independent diagnosis and is also higher than that of the AFP independent diagnosis, that is, the TIGAR in the blood plasma can be independently or jointly used as a molecular marker for diagnosing the recurrence of the liver cancer.

Description

technical field [0001] The invention belongs to the technical field of biotechnology detection, and specifically relates to the application of TIGAR protein in blood plasma as a molecular marker in the preparation of a kit for predicting and diagnosing liver cancer recurrence. Background technique [0002] Cancer is a heterogeneous disease with high morbidity, high mortality and poor prognosis. Worldwide, more people die from cancer than from other communicable diseases such as AIDS, malaria and tuberculosis combined. Primary liver cancer (PHC) is a very common tumor with high malignancy, high recurrence rate and high mortality rate. In the statistics of the most common cancer in the world, liver cancer ranks sixth; and it ranks the third among the causes of cancer death Four, about 841,000 new cases (4.7%) and 782,000 deaths per year (8.2%); it is also the fifth most common cancer in men (523,000 cases, accounting for 7.9% of the total) and the second most common cancer T...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/577G01N33/573G01N21/76
CPCG01N33/57438G01N33/57488G01N33/57476G01N33/577G01N33/573G01N21/76G01N2333/916
Inventor 刘素嬛张婷李滨
Owner THE FIRST AFFILIATED HOSPITAL OF XIAMEN UNIV