Isolate the test samples soon after collecting, then, analyze immediately (within 2 hours). Or aliquot and store at -20°C for long term. Avoid multiple freeze-thaw cycles.
Cell lysate sample Collection and Storage
When you get the cell by cell culture centrifugation, add the lysis buffer to the cell (Should be strictly according to the lysis buffer manual, it is easy to buy the lysis buffer in market), then mix well, after 5 minutes’ standing, you can get the sample for detection.
Cell culture supernate
Centrifuge supernate for 20 minutes to remove insoluble impurity and cell debris at 1000×g at 2 – 8°C. Collect the clear supernate and carry out the assay immediately.
Allow samples to clot for 2 hours at room temperature or overnight at 4°C before centrifugation for 20 minutes at approximately 1000×g. Collect the supernatant and carry out the assay immediately. Blood collection tubes should be disposable, non-pyrogenic, and non-endotoxin.
Collect plasma using EDTA-Na2 as an anticoagulant. Centrifuge samples for 15 minutes at 1000×g at 2 – 8°C within 30 minutes of collection. Collect the supernatant and carry out the assay immediately. Avoid hemolysis, high cholesterol samples.
For general information, hemolysis blood may affect the result, so you should rinse the tissues with ice-cold PBS (0.01M, pH=7.4) to remove excess blood thoroughly. Tissue pieces should be weighed and then minced to small pieces which will be homogenized in PBS (the volume depends on the weight of the tissue. 9mL PBS would be appropriate to 1 gram tissue pieces. Some protease inhibitor is recommended to add into the PBS.) with a glass homogenizer on ice. To further break the cells, you can sonicate the suspension with an ultrasonic cell disrupter or subject it to freeze-thaw cycles. The homogenates are then centrifugated for 5minutes at 5000×g to get the supernate.
Other biological fluids
Centrifuge samples for 20 minutes at 1000×g at 2 – 8°C. Collect the supernatant and carry out the assay immediately.