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Human bladder cancer cellular chemiluminescent detection kit and preparation method thereof

A technology of bladder cancer cells and a kit is applied in the fields of immunoassay and clinical diagnosis medicine, and achieves the effect of short detection time and easy operation.

Active Publication Date: 2013-04-10
INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] This invention focuses on the early diagnosis of bladder cancer and the follow-up observation of bladder cancer patients, and intends to solve the current difficult problem in the diagnosis of bladder cancer, that is, to use two specific monoclonal antibodies to identify bladder cancer cells to realize double antibody recognition of bladder cancer cells And capture, combined with highly sensitive chemiluminescence immunoassay method, quantitative detection of bladder cancer cells exfoliated in urine, does not require expensive detection equipment, is convenient for clinical promotion, has good market application value, and will make up for the shortage of existing kits in the market

Method used

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  • Human bladder cancer cellular chemiluminescent detection kit and preparation method thereof
  • Human bladder cancer cellular chemiluminescent detection kit and preparation method thereof
  • Human bladder cancer cellular chemiluminescent detection kit and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Example 1 Preparation of a new bladder cancer cell chemiluminescence detection kit of the present invention

[0057] 1. Preparation of standardized bladder cancer cells

[0058] Bladder cancer EJ cell line (CRL-2888 TM ) were purchased from the American Type Culture Collection (ATCC), and cultured in DMEM medium (Gibco, 12100-046) containing 10% fetal bovine serum. Add 1-2 mL of trypsin (Invitrogen, R-001-100) to the cultured cells with good adherence, digest at 37°C for 2 minutes, add an appropriate amount of medium, centrifuge the digested cells, discard the supernatant, and resuspend in Fetal bovine serum plus 10% dimethyl sulfoxide (DMSO) prepared freezing solution, with 10 6 The concentration of cells / mL is frozen at -80°C, and each tube is frozen in 500 μL. During the transportation of the kit, this component needs to be stored and transported separately in dry ice.

[0059] 2. Preparation of monoclonal antibody BCMab2 hybridoma cells

[0060] Take frozen EJ ...

Embodiment 2

[0094] Example 2 Method of using the bladder cancer urine exfoliated cytochemical method photoimmunoassay assay kit of the present invention

[0095] 1. Sample pretreatment

[0096] Take the first morning urine or the second morning urine sample. If it is tested immediately, the sample does not need to be processed and can be tested directly. If the test cannot be performed on the same day, the sample needs to be frozen: take 10 mL of urine, centrifuge at 3000 rpm for 5 minutes, suspend the urine sediment in 500 uL of cell freezing solution (fetal bovine serum containing 10% DMSO), place it at 4°C for 2 hours, and then Store at -80°C for later use.

[0097] 2. Detection method

[0098] Before using this kit for experiments, first adjust the incubator or water bath to 37°C; then take out the magnetic particle solution, HRP-labeled antibody and each buffer solution prepared in Example 1, and place them at room temperature to equilibrate to room temperature; then take out the ...

Embodiment 3

[0105] The methodological index of embodiment 3 kits of the present invention

[0106] The test kit prepared in Example 1 is tested according to the conventional manufacturing and testing procedures in the art, and the results are as follows:

[0107] 1. Determination of kit precision

[0108] (1) Precision experiment of EJ cell standard

[0109] Extract 10 kits from the kit prepared in Example 1, and measure 1×10 according to the operation described in Example 2 3 Each / mL EJ cell standard 5 times. The coefficient of variation of the measurement results was calculated, and the measurement results are shown in Table 1. The results showed that the coefficient of variation was between 3.5% and 10%.

[0110] Table 1 EJ Cell Standard Reproducibility Experiment

[0111]

[0112] (2) Sample precision experiment

[0113] 10 kits were extracted from the kits prepared in Example 1, and the urine exfoliated cells of two bladder cancer patients were measured according to the opera...

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Abstract

The invention relates to the field of immune analysis medicine, and particularly provides a kit for detecting human bladder cancer by a chemiluminescent analysis method on the basis of immune magnetic particles and a preparation method for the kit. According to the invention, the kit mainly comprises: 1) a bladder cancer EJ cell standard product; 2) magnetic particles which are coated by monoclonal antibodies capable of specifically identifying bladder cancer cell surface antigen; 3) enzyme-labeled monoclonal antibodies capable of specifically identifying bladder cancer cell surface antigen; 4) a chemiluminescent substrate which the enzyme in the 3) acts on; 5) a reaction tube or a microporous plate; and 6) a magnetic separator matched with the reaction tube or the microporous plate in the 5).

Description

technical field [0001] The present invention relates to the fields of immune analysis and clinical diagnostic medicine, specifically, the present invention provides a chemiluminescent detection kit for human bladder cancer cells and a preparation method thereof. Background technique [0002] Bladder cancer is the most common tumor in the urinary system. Hematuria is one of the early clinical symptoms of bladder cancer patients, including gross hematuria and microscopic hematuria. However, hematuria also occurs in urinary tract inflammation and stone symptoms, and is not a specific symptom of bladder cancer. It is precisely because the early symptoms of bladder cancer are not obvious that hematuria is found, and bladder tumors in patients generally develop into larger tumor nodules. Finding a specific diagnostic method for bladder cancer has important clinical application value for the diagnosis of early bladder cancer. [0003] There are several methods for early diagnosi...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/531
Inventor 范祖森张倩云李翀杜颖王彦英杨昭
Owner INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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