Low-temperature dissociation kit suitable for single cell sequencing and application thereof
A single-cell sequencing and kit technology, which is applied in the biological field, can solve the problems that protease cannot be stored for a long time and limit its wide application, and achieve the effects of simple operation, improving data quality and avoiding operation errors.
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Embodiment 1
[0083] This example describes the application of the kit of the present invention (stored at -20°C for 6 months) in single-cell sequencing of rat lung tissue. The working concentration of low-temperature protease in the kit of the present invention is 5 mg / mL, and the CaCl in enzymolysis buffer 1 2 The working concentration of DNase I was 5 mM, and the working concentration of DNase I in enzymatic digestion buffer 2 was 100 U / mL.
[0084] 1) Prepare reagents.
[0085] Before starting to prepare samples, take out the enzymatic hydrolysis solution, enzymatic hydrolysis buffer 1, and enzymatic hydrolysis buffer 2 from the -20°C refrigerator, and place them on ice until they melt.
[0086] 2) Prepare the sample.
[0087] The freshly obtained rat lung tissue was placed in a petri dish containing DMEM medium pre-cooled on ice. Rinse the tissue with pre-cooled DMEM medium on ice, suck out the rinsed medium, and then add new pre-cooled medium, repeat this process 3 times until no o...
Embodiment 2
[0105] This example describes the application of the kit of the present invention (-20°C, stored for 6 months) in single-cell sequencing of human tonsil tissue. The working concentration of low-temperature protease in the kit of the present invention is 5 mg / mL, and the CaCl in enzymolysis buffer 1 2 The working concentration of DNase I was 5 mM, and the working concentration of DNase I in enzymatic digestion buffer 2 was 100 U / mL.
[0106] 1) Prepare reagents.
[0107] Before starting to prepare samples, take out the enzymatic hydrolysis solution, enzymatic hydrolysis buffer 1, and enzymatic hydrolysis buffer 2 from the -20°C refrigerator, and place them on ice until they melt.
[0108] 2) Prepare the sample.
[0109] The freshly obtained human tonsil tissue was placed in a petri dish containing DMEM medium pre-cooled on ice. Place the petri dish on ice, rinse the tissue with DMEM medium pre-cooled on ice, suck out the rinsed medium, and then add new pre-cooled medium, repea...
Embodiment 3
[0127] This example describes the application of the kit of the present invention (stored at -20°C for 6 months) in single-cell sequencing of mouse liver tissue. The working concentration of low-temperature protease in the kit of the present invention is 5 mg / mL, and the CaCl in enzymolysis buffer 1 2 The working concentration of DNase I was 5 mM, and the working concentration of DNase I in enzymatic digestion buffer 2 was 100 U / mL.
[0128] 1) Prepare reagents.
[0129] Before starting to prepare samples, take out the enzymatic hydrolysis solution, enzymatic hydrolysis buffer 1, and enzymatic hydrolysis buffer 2 from the -20°C refrigerator, and place them on ice until they melt.
[0130] 2) Prepare the sample.
[0131] The freshly harvested mouse liver tissue was placed in a petri dish containing DMEM medium pre-cooled on ice. Place the petri dish on ice, rinse the tissue with DMEM medium pre-cooled on ice, suck out the rinsed medium, and then add new pre-cooled medium, re...
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