Target MLLT11 for preventing lung cancer and application thereof
A lung cancer and target technology, applied in the biological field, to achieve the effect of inhibiting malignant transformation and preventing lung cancer
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Embodiment 1
[0046] Example 1. Normal lung bronchial epithelial cells were treated with arsenic, and the protein expression level of MLLT11 was up-regulated in a concentration- and time-dependent manner
[0047] 1. Cell infection treatment
[0048] (1) Treatment of acute arsenic exposure of cells:
[0049] a) Count the cells of human normal bronchial epithelial cells BEAS-2B. A total of 200,000 cells are spread in each well of a six-well plate, with a total of 8 wells. Using DMEM medium containing 10% FBS, keep the temperature at 37°C in CO 2 The culture was carried out in an incubator; the cell density reached about 80% after 12 hours.
[0050] b) 12 hours after seeding the plate, the original DMEM medium containing 10% FBS was replaced with 0.1% FBS DMEM medium, and the cells were starved for 12 hours according to the same culture conditions as above.
[0051] c) After the cells were starved for 12 hours, two groups of experiments were carried out, each with 4 wells of cells. One grou...
Embodiment 2
[0071] Example 2. The expression level of MLLT11 in clinical lung cancer tissues generally shows a high trend.
[0072] 1. Clinical tissue immunohistochemistry
[0073] (1) Paraffin-embedded sections:
[0074] a) Tissue collection: Lung cancer tissue and its corresponding paracancerous tissue were collected. This operation strictly abides by the relevant regulations of the Medical Ethics Committee of Wenzhou Medical University.
[0075] b) Tissue fixation: lung cancer tissue was fixed with 4% PFA, and fixed at 4°C for 24-48h.
[0076] c) Tissue dehydration: the fixed lung cancer tissue was washed with running water, and the PFA was cleaned. Then put the specimen in 30% ethanol for 30min, 50% ethanol for 30min, 70% ethanol for 1h, 80% ethanol for 1h, 85% ethanol for 1h, 95% ethanol for 1h, 95% ethanol for 1h, 100% ethanol for 1h, 100% ethanol for 1h gradient dehydration.
[0077] d) Tissue transparency: After the tissue is dehydrated, put the tissue in a solution mixed wit...
Embodiment 3
[0101] Example 3, MLLT11 significantly promotes the colony formation of BEAS-2B cells induced by arsenide for a long time.
[0102] 1. Steps related to cell transfection (1) Lipofectamine transfection method:
[0103] a) BEAS-2B cells were plated for 24 hours, and when the cells grew to 80%, the medium was replaced 1 hour in advance with fresh 10% FBSDMEM+++ medium.
[0104] b) Add 2 μg of MLLT11 KO plasmid (Vector, KO#1, KO#2, KO#3) into 50 μL of serum-free DMEM containing high glucose and mix thoroughly.
[0105] c) Take 6μL PolyJet TM Add 50 μL of serum-free DMEM containing high glucose to the reagent, and mix gently with a gun for 3-4 times.
[0106] d) Immediately place the diluted PolyJet TM The reagent was added to the diluted DNA solution, and mixed up and down gently for 3-4 times. (sequence only PolyJet TM plus DNA)
[0107] e) Incubate at room temperature for 15 minutes, not exceeding 20 minutes.
[0108] f) Dispense 100μL PolyJet TM Add the DNA mixture drop...
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