4A7G7B6 reacts with CD25 antigen, a chain of low-affinity interleukin-2 receptor ( IL-2Ra ), which is expressed on activated cells including T, B, NK cells and monocytes.
Optimal working dilution should be determined by the investigator.
Testdurchführung
Take 100 μL peripheral blood anticoagulated by EDTA and add to the bottom of 5 mLtube,
Add appropriate amount of antibody to the bottom of flow tube mixing with the whole blood, incubate for 30 minutes at room temperature,
Add 2 ml1×RBC lysis buffer, incubate for 10 minutes 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, then 1000 rpm centrifugation for 5 minutes, discard the supernatant,
Add appropriate amount of fluorescent-labeled Streptavidin and incubate for 20 minutes away from light at room temperature.
Repeat step 5.
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).
The antigen also prsent on subset of thymocytes, HTLV-1 transformed T cell lines, EBV transformed B cells, myeloid precursors and oligodendrocytes. The high affinity IL-2 receptor is formed by the noncovalent association of of a ( 55 kDa, CD25 ), b ( 75 kDa , CD122 ), and g subunit ( 70 kDa, CD132 ). The interaction of IL-2 with IL-2R induces the activation and proliferation of T, B , NK cells and macrophages. CD4+/CD25+ cells might directly regulate the function of responsive T cells.