Application of SNEP1 protein in diagnosis of colorectal cancer
A colorectal cancer and protein technology, applied in the application field of SNEP1 protein in the diagnosis of colorectal cancer, can solve the problems of no collection, no functional description, etc.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0027] Example 1: Plate cloning method to detect the effect of SNEP1 on the proliferation ability of tumor cells
[0028] Take SNEP1 overexpression or silencing stable tumor cells or control tumor cells in the logarithmic growth phase, remove the medium, add trypsin to fully digest for 2-3min, then add 500 μl DMEM high-glucose medium containing 10% serum to neutralize the trypsin, Mix the cells by pipetting to separate the cells into single cells. Seed 1000 cells / well into a 6-well plate and disperse the cells evenly. Place at 37°C, 5% CO 2 Culture in the incubator for 14-20 days. Culture was terminated when macroscopic colonies were observed. Remove the medium, add 4% paraformaldehyde solution and fix at 37°C for 10min. After removing the fixative, wash with PBS 3 times, add 0.2% crystal violet and stain for 20-30min. The staining solution was washed away with PBS several times until the PBS was clear and then air-dried. Place the 6-well plate upside down on a professio...
Embodiment 2
[0029] Example 2: Immunohistochemical detection of SNEP1 expression in cancer tissue and normal tissue
[0030] The resected tissue samples were placed in a sterile tray, and tissue scissors and scalpels were used to collect the tissue pieces, and the tissue blocks were fixed in 10% neutral formalin for 48 hours. The fixed tissue block was trimmed slightly, put in the embedding box, passed through 30%, 50%, 70%, 80%, 90%, 95%, 100% ethanol solution and xylene I, xylene II in sequence, Finally, paraffin I and paraffin II were added to dehydrate the tissue gradient. Then start the paraffin embedding machine to embed the tissue block. Use a paraffin trimmer to cut the tissue block into 3 μm / slice slices, hang the slices on a glass slide, spread them out, and place them on a 65°C oven for 30 minutes. After the paraffin sections were baked in a 70°C oven for 90 minutes, they were dewaxed and hydrated by xylene I, xylene II, and 100%, 95%, and 85% ethanol solutions in sequence. T...
Embodiment 3
[0031] Example 3: Western blot detection of SNEP1 expression in cancer tissue and normal tissue Prepare crushed ice, weigh and mark the 5ml centrifuge tube, and pre-cool it on ice with PBS; Cup transfer, cut part of the tissue, put it into a pre-cooled centrifuge tube, put the remaining tissue back into the cryovial, and put it back to the original place; weigh the tissue and record; wash the tissue with pre-cooled PBS 2-3 times, wash De-adhered blood; cut up the tissue, add RIPA buffer containing 1:100 protease inhibitor and 1mMPMSF, add the ratio tissue: buffer=1:10 (g / ml); homogenize the tissue on a tissue disruptor, 13000g, 10s until the tissue was minced; the tissue should be kept on ice at all times; on a rotator, rotate at 4°C for 30 minutes to fully lyse the tissue; transfer the lysate to an EP tube, centrifuge at 4°C, 12000g, 15min; take the supernatant, Obtain the required tissue protein lysate; take 100 μl of protein for quantification and do WB, and store the rest ...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More