Fluorescent visualization and quantitative detection of telomerase activity based on water-soluble cationic polymer
A water-soluble cation, quantitative detection technology, applied in the field of biosensing, can solve the problems of complex operation, high cost, low sensitivity, etc., and achieve the effects of simple experimental operation, rapid response, and high sensitivity
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Embodiment 1
[0039] A method for quantitatively detecting telomerase activity based on fluorescence visualization of a water-soluble cationic polymer, comprising the following steps:
[0040] 1) Cell culture and telomerase extraction to obtain lysed cells: HeLa cells were inoculated in DMEM medium containing 10% fetal bovine serum, penicillin and streptomycin, and incubated in 5% CO 2 , cultured in a 37°C incubator. All cells were collected during exponential growth phase. Aliquot 1 million cells into 1.5 mL EP tubes, wash twice with ice-cold phosphate buffered saline (pH=7.4), and redisperse in 200 μL of ice-cold CHAPS lysis buffer (10 mM Tris-HCl, pH 7.5, 1mM MgCl 2 , 1 mM EGTA (ethylene glycol bis(2-aminoethyl ether) tetraacetic acid), 0.5% (w / v) CHAPS (3-[3-(cholamidopropyl)dimethylamino]propanesulfonic acid inner salt ), 10% (v / v) glycerol (glycerol), 0.1 mM PMSF (phenylmethylsulfonyl fluoride)). Cells were incubated on ice for 30 minutes and then centrifuged at 12000 rpm for 20 m...
Embodiment 2
[0044] Embodiment 2: A method for quantitatively detecting telomerase activity based on fluorescence visualization of water-soluble cationic polymers, comprising the following steps:
[0045] 1) Cell culture and telomerase extraction to obtain lysed cells: HeLa cells were inoculated in DMEM medium containing 10% fetal bovine serum, penicillin and streptomycin, and incubated in 5% CO 2 , cultured in a 37°C incubator. All cells were collected during exponential growth phase. Aliquot 1 million cells into 1.5 mL EP tubes, wash twice with ice-cold phosphate buffered saline (pH=7.4), and redisperse in 200 μL of ice-cold CHAPS lysis buffer (10 mM Tris-HCl, pH 7.5, 1mM MgCl 2 , 1 mM EGTA, 0.5% (w / v) CHAPS, 10% (v / v) glycerol, 0.1 mM PMSF). Cells were incubated on ice for 30 minutes and then centrifuged at 12000 rpm for 20 minutes at 4°C. After centrifugation, the clarified lysate was carefully transferred to a new EP tube to obtain the lysed HeLa cells, which were quickly frozen a...
Embodiment 3
[0050] A method for quantitatively detecting telomerase activity based on fluorescence visualization of a water-soluble cationic polymer, comprising the following steps:
[0051]1) Cell culture and telomerase extraction to obtain lysed cells: HeLa cells were inoculated in DMEM medium containing 10% fetal bovine serum, penicillin and streptomycin, and incubated in 5% CO 2 , cultured in a 37°C incubator. All cells were collected during exponential growth phase. Aliquot 1 million cells into 1.5 mL EP tubes, wash twice with ice-cold phosphate buffered saline (pH=7.4), and redisperse in 200 μL of ice-cold CHAPS lysis buffer (10 mM Tris-HCl, pH 7.5, 1mM MgCl 2 , 1 mM EGTA, 0.5% (w / v) CHAPS, 10% (v / v) glycerol, 0.1 mM PMSF). Cells were incubated on ice for 30 minutes and then centrifuged at 12000 rpm for 20 minutes at 4°C. After centrifugation, the clarified lysate was carefully transferred to a new EP tube to obtain the lysed HeLa cells, which were quickly frozen and stored in a...
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