The streptavidin fluorochrome conjugates are commonly used in indirect staining protocols to detect biotinylated primary antibodies in flow cytometry. Streptavidin binds to biotin with high affinity. Streptavidin-PE has been reported for use in flow cytometric analysis, immunohistochemical staining, and immunocytochemistry.
Optimal working dilution should be determined by the investigator.
实验流程
Take 100 μL peripheral blood anticoagulated by EDTA and add to the bottom of 5 mLtube,
Add biotinylated primary antibody to the bottom of flow tube mixing with the whole blood, incubate for 20 minutes at room temperature away from light,
Add 2 ml1×RBC lysis buffer, incubate for 10 minutes away from light after mixing, dissolve red blood cells (recommended: RBC lysing Solution 10×,Cat.: FXP001),
Sample tube is set to 1000 rpm centrifugation for 5 minutes, discard the supernatant,
Add 2 mLPBS wash buffer to resuspend the cells, then1000 rpm centrifugation for 5 minutes, discard the supernatant,
Add 10 μL Streptavidin-FITC mixing with cells, incubate for 20 minutes at room temperature away from light,
Add 2 mLPBS wash buffer to resuspend the cells, then1000 rpm centrifugation for 5 minutes, discard the supernatant,
Add 0.5 mLPBS wash buffer to resuspend the cells and detect by flow cytometry (sample should be determined on the day on the machine and can also be added fixation overnight at 4 °C then measured).