PMC:1315352 / 15504-16445 JSONTXT

{"project":"CellFinder","denotations":[{"id":"T267","span":{"begin":597,"end":602},"obj":"Species"},{"id":"T268","span":{"begin":603,"end":606},"obj":"GeneProtein"}],"text":"Flow cytometry\nAntibodies were prepared at the concentration of 0.1 mg/mL. 10 μL of the stock solution was added to 1 – 2.5 × 105 cells in a total reaction volume not exceeding 200 μL. The sample was then incubated for 20 min at 2–8 °C. Following incubation, excess antibody was removed by washing cells twice with FACS buffer (2% FCS and 0.1% sodium azide in Hank's buffer). After wash, cells were resuspend in 200 μL of FACS buffer and the binding of unlabeled monoclonal antibodies was visualized by adding 10 μL of a 25 μg/mL stock solution of a secondary developing reagent such as goat anti-mouse IgG conjugated to a fluorochrome for 20 min at 2–8°C. Following incubation, cells were washed once with FACS buffer, once with PBS. After wash, cells were resuspend in 400 μL of PBS and analyzed on a FACScant flow cytometer (Becton-Dickinson, Mountain View, CA). Five thousand events were collected and analyzed using CELL Quest software."}