Preparation of cell and cell nucleus microarray
A cell nucleus and microarray technology, applied in biochemical equipment and methods, microbial measurement/inspection, etc., can solve problems such as complex procedures, uneven cell density, cell fragmentation, etc., and achieve simple production process, strong operability, The effect of large array capacity
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Embodiment 1
[0043] 1. Treatment of cultured cells: Adherent cells, human nasopharyngeal carcinoma CNE1, CNE2, HNE1, HNE2, 5-8F and 6-10B nasopharyngeal carcinoma cell lines and 6-10B cells transfected with KIAA1173 gene, A total of 7 cell lines were cultured. First, the cultured cells were digested with 0.25% trypsin, washed three times with phosphate buffer, centrifuged at 1500 rpm for 5 min each time, and then washed with 1:3 (volume ratio) methanol and glacial acetic acid solution for 15 min, and adjust the concentration of the cell suspension to 1×10 4 / μl for use.
[0044] 2. Fabrication of the microarray wax film: first make a blank wax block, and then punch holes in the blank wax block with a 0.6mm needle of the tissue microarray instrument, and the number of holes is 7×7. Freeze the prepared wax blocks with holes at -20°C for 30 minutes; cut into 20 μm thick slices, stick them on siliconized glass slides, and bake in an oven at 60°C for 10 seconds.
[0045] 3. Production of cell...
Embodiment 2
[0049] 1. Extract nuclei from paraffin tissue. 60 cases of diffuse large B-cell lymphoma were first stained with HE to observe the distribution of tumor cells, sliced (three pieces in total), and the thickness was 10 microns; put the wax slices into a 1.5ml polypropylene centrifuge tube, add xylene 1ml, shake slightly for 10min, slowly pour out the xylene solution, add fresh xylene and shake gently for 10min; discard the xylene, add 1ml ethanol and shake gently for 10min, pour out the ethanol, replace with fresh ethanol and shake again for 10min, 20ml of 80% ethanol, Dehydrate 1ml of 70% ethanol step by step, 10min each time, discard the ethanol, add 50ml of water and centrifuge to form a loose tissue precipitate; add 1ml of proteinase K digestion solution (0.1mg / ml proteinase K and 0.5% SDS in TE ), incubated at 37°C with slight shaking, and digested for 2 hours; after the mixture was shaken sufficiently, it was centrifuged at 1000 rpm for 5 sec, and the supernatant was the...
Embodiment 3
[0055] 1. Treatment of cultured cells: Cells growing in suspension, three lymphoma cell lines, L428, DG75 and jurkit, were cultured, cells in good growth condition were selected, washed with phosphate buffer three times, each time at 1500 rpm , centrifuged for 5 min, then fixed with 1:3 methanol and glacial acetic acid solution for 15 min, and adjusted the concentration of the cell suspension to 1×10 4 / μl for use.
[0056] 2. Fabrication of the microarray wax film: first make a blank wax block, and then punch holes in the blank wax block with a 1.0 mm needle of the tissue microarray instrument, the number of holes is 4×4. Freeze the prepared wax blocks with holes at -10°C for 15 minutes; cut into 20 μm thick slices, paste them on siliconized glass slides, and bake in an oven at 60°C for 5 sec.
[0057] 3. Production of cell microarray: Inject the cell suspension obtained in step 1 into the wells in the paraffin film respectively, inject 5 or 6 wells for each cell line, a tot...
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