Method of quickly separating breast cancer primary tumor living cell
A breast cancer and living cell technology, applied in the field of cell separation, can solve the problems of fibroblast contamination, low purity of separation of breast cancer cells, inability to use clinical drug sensitivity tests, etc., and achieve the effect of broad material sources
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Embodiment 1
[0058] Embodiment 1 sample collection
[0059] All samples used in the implementation of the present invention are derived from surgical resection specimens of breast cancer patients, and all participating patients have given informed consent to the scientific research.
[0060] The detection samples used in the present invention are usually collected in a clinically acceptable manner, preferably in a manner in which nucleic acid (especially RNA) or protein is protected.
[0061] Patient information (age, gender, impact reports, course of treatment, other medical conditions, family history, etc.) was obtained from the hospital database and used to match the various samples collected. Pathological follow-up studies (such as histological analysis by hematoxylin and eosin staining, ie H&E staining) are used to clarify the disease status of a given sample and to ensure consistent grading of samples.
[0062] Fresh breast cancer specimens were obtained in the operating room and se...
Embodiment 2
[0063] Embodiment 2: tissue pretreatment
[0064] The pathologist cut out a 1cm3 tissue block from the non-necrotic area of the sample to be tested and the area containing more tumor tissue. At the same time, the tissue block was subjected to frozen section and HE staining for quality control to ensure that the tumor cell content reached more than 75%. Place the tissue in 10ml of 199 culture medium containing gentamicin, and store it on ice for 24 hours for specimen transportation.
Embodiment 3
[0065] Example 3: Rapid batch separation of high-purity primary breast cancer cells
[0066] 1cm 3 Large and small breast cancer tissue blocks cut into 1mm 3 For tissue fragments of different sizes, wash the tissue twice with 10ml of 199 culture medium containing gentamicin, and repeatedly pipette for 2 minutes, transfer 10ml of 199 culture medium containing tissue to a 15ml centrifuge tube and let it stand for 1 minute, carefully Remove the supernatant to another clean 15ml centrifuge tube, and avoid aspirating the tissue pieces at the bottom, centrifuge the supernatant obtained above at a speed of 1000RPM for 5 minutes, discard the supernatant carefully, and use 10ml of Ham'sF-12 Resuspend the cell pellet in the culture medium, in which Ham'sF-12 culture medium contains 5% FBS, 1 μg / ml insulin and 1 μg / ml hydrocortisone, move the obtained cells to a culture dish, at 37oC, 9% CO 2 cultured in an environment, observe the obtained cells with a microscope, count and take pi...
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