Method of stable labelling liver cancer cell

A technology for liver cancer cells and human liver cancer cell lines, applied in the field of stable labeling of liver cancer cells, can solve the problems of high cost, complicated operation, and poor practicability, and achieve the effect of convenient operation, simplified steps, and strong practicability

Inactive Publication Date: 2006-06-28
WUHAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

QD currently adopted abroad s The method of labeling cells is mainly through the combination of streptavidin-biotin system. This method is relatively complicated to operate and has high cost

Method used

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  • Method of stable labelling liver cancer cell
  • Method of stable labelling liver cancer cell
  • Method of stable labelling liver cancer cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] The present invention is to the fluorescent labeling of human liver cancer cell HCCLM6, and the steps are as follows:

[0035] A. Cell culture The human liver cancer cell line HCCLM6 was cultured in RPMI-1640 medium (Sigma, Saint Louis, Missouri, USA) containing 10% fetal bovine serum (Hyclone, Utah, USA). Culture conditions: 37°C, 5% CO 2 / 95% air, saturated humidity. Replace the culture medium every other day, and subculture once every 4-6 days.

[0036] B. Treatment of cell culture slides and culture dishes, wash the 20mm×20mm cover glass and the glass culture dish with a diameter of 9cm with clean water, soak in 2% (vol / vol) hydrochloric acid solution overnight, soak in clean water, and use a soft brush Gently scrub, dry in an oven, control the temperature at 50-70°C, soak in 100ml of concentrated sulfuric acid, potassium dichromate cleaning solution, and 1000ml of distilled water in the acid tank for 1-3 days, rinse with tap water 20 times, wash with distilled wa...

Embodiment 2

[0050] The specificity test of the marker liver cancer cells of the present invention, the steps are as follows:

[0051] A.~G. The same as in Example 1, divide the sealed hepatoma cell slides into two groups, one group uses specific mouse anti-human AFP antibody as the primary antibody, and one group uses PBS (same as in Example 1, step D) as the primary antibody. Primary antibody, according to the steps of Example 1

[0052] H. Operation.

[0053] I.~K. are the same as embodiment 1

[0054] L. The slides were mounted in buffered glycerol and observed under a confocal laser scanning fluorescence microscope. see results figure 2 .

Embodiment 3

[0056] The fluorescence stability test of labeled liver cancer cells of the present invention, the steps are as follows:

[0057] A.~I. Same as Example 1, the liver cancer cell slices that have been washed are divided into two groups, and one group is divided into QD s -IgG complex probe was used as the secondary antibody, and one group used FITC-IgG as the secondary antibody, and operated according to step J. of Example 1.

[0058] K. same as embodiment 1

[0059] L. The slides were mounted in buffered glycerol and observed under a confocal laser scanning fluorescence microscope. see results image 3 , Figure 4 and Figure 5 .

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Abstract

A method for stably labeling liver cancer cell includes combining liver cancer cell with specific AFP single cloning antibody used as the first resistance factor, washing and climbing plate of PBS, carrying out fluorescent label by using QDs ¿C IgG composite probe as the second resistance factor, washing and climbing plate of PBS, re ¿C staining cell core with DAPI, washing and climbing plate of PBS and preserving sample in light shielded condition.

Description

technical field [0001] The invention belongs to the technical field of quantum dot immunofluorescence cytochemistry, and in particular relates to a method for stably marking liver cancer cells. Background technique [0002] Immunofluorescence cytochemistry uses known antibodies (or antigens) labeled with fluorescent substances as probes to detect target antigens (or antibodies) in tissues and cell samples to be tested, and the antigen-antibody complexes formed have fluorescent substances on them. Under the fluorescence microscope, due to the excitation light source, the fluorescent substance emits bright fluorescence, so that the location and nature of the antigen (or antibody) can be distinguished. This type of technology is widely used in modern biology and medicine. As early as more than 60 years ago, the immunofluorescence technology created by Coons et al. has made great progress. With the continuous development of high-sensitivity and high-s...

Claims

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Application Information

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IPC IPC(8): G01N33/532G01N33/574G01N33/577
Inventor 陈良冬李雁刘佳庞代文
Owner WUHAN UNIV
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