| Product Description |
PMSF (ST506) is required, but not supplied in this product. Inhibitor cocktails can also be used for better lysis results. provides a variety of inhibitor cocktails, such as Protease and Phosphatase Inhibitor Cocktail for General Use , Protease and Phosphatase Inhibitor Cocktail for General Use, MS-safe , Protease and Phosphatase Inhibitor Cocktail for Mammals , Protease and Phosphatase Inhibitor Cocktail for plant , Protease and Phosphatase Inhibitor Cocktail for Fungi , and Protease and Phosphatase Inhibitor Cocktail for Bacteria . If phosphorylated proteins are not to be detected, inhibitors of protease only can be used.All procedures for sample lysis should be performed on ice or at 4℃.
An appropriate sample lysis buffer can be selected by referring to the website at https://www.aladdin-e.com or optimized by preliminary tests.
This product is for R&D only. Not for drug, household, or other uses.For your safety and health, please wear a lab coat and disposable gloves during the operation.
Instructions for Use：
1. Lysis of cultured cells
a. Thaw Plant RIPA Lysis Buffer completely and mix well. Just before use, take an appropriate amount of RIPA Lysis Buffer and add PMSF to a final concentration of 1mM. Other protease and phosphatase inhibitors can also be used if required.
b. Collect cells by centrifugation, add 100-200μl of lysis buffer per 0.5-1.0×106 cells. Pipette to resuspend cells and lyse cells on ice for 2-10 min.
c. After full lysis of cells, centrifuge at 10,000-14,000×g for 3-5 minutes and take the supernatant for PAGE, Western or IP analysis.
2. Lysis of protoplasts
a. Cut tissues into small pieces and prepare protoplasts.
b. (Optional) Transform the plasmid into protoplasts and continue incubation for 16-48 hours. Treat as desired.
c. Centrifuge at 100-500×g to collect protoplasts.
d. Thaw Plant RIPA Lysis Buffer completely and mix well. Just before use, take an appropriate amount of Lysis Buffer and add PMSF to a final concentration of 1mM. Other protease and phosphatase inhibitors can also be used if required.
e. Add 100-200μl of lysis buffer per 0.5-1.0×106 protoplasts, flick the bottom of the tube to lyse thoroughly. There should be no obvious precipitates after full lysis. For large amounts of protoplasts, dispense them into 0.5-1.0×106 protoplasts per tube for lysis.
3. Lysis of tissues
a. Cut tissues into small pieces.
b. Thaw Plant RIPA Lysis Buffer completely and mix well. Just before use, take an appropriate amount of Lysis Buffer and add PMSF to a final concentration of 1mM. Other protease and phosphatase inhibitors can also be used if required.
c. Add 100-200μl of lysis buffer per 20mg of tissues. The amount of lysis buffer can be adjusted based on the lysis results.
d. Homogenize tissues thoroughly with a glass homogenizer or 's TissueMasterTM Handheld Homogenizer .
e. Centrifuge at 10,000-14,000×g for 3-5 minutes and take the supernatant for PAGE, Western or IP analysis.
f. For tiny tissues, add lysis buffer after appropriate cutting and lyse tissues with vigorous vortex. Centrifuge and take the supernatant for subsequent analysis. This lysis method is convenient, with no homogenization needed, but the lysis result is not as thorough as that from the homogenization method.
Note: It is normal to see a small clump of transparent gelatinous substances containing genomic DNA in RIPA cell lysate. In cases where DNA-binding proteins will not be analyzed, the supernatant can be analyzed directly. Otherwise, break the gelatinous substance with an ultrasonicator and analyze the supernatant after centrifugation. Some common transcription factors such as NF-kappaB and p53 can be detected in the supernatant without ultrasonic processing
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