APC Mouse Anti-Human CD71

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Incubation 60 minutes at 4°C in staining buffer (2% BSA in PBS). Corresponding isotypes were used as control. Cells were washed in PBS, centrifuged, and finally resuspended in 300-μL analysis buffer (1% BSA/2 mmol/L EDTA in PBS).	The dead cells and debris were excluded after 7-AAD staining (Invitrogen). Data were collected on FACSAriaIII cell sorter (BD Biosciences) and analyzed using FlowJo software (TreeStar).

Protocol tips
Incubation 60 minutes at 4°C in staining buffer (2% BSA in PBS). Corresponding isotypes were used as control. Cells were washed in PBS, centrifuged, and finally resuspended in 300-μL analysis buffer (1% BSA/2 mmol/L EDTA in PBS).
Downstream tips
The dead cells and debris were excluded after 7-AAD staining (Invitrogen). Data were collected on FACSAriaIII cell sorter (BD Biosciences) and analyzed using FlowJo software (TreeStar).

Publication protocol

For flow cytometry analysis, cell suspension (1 × 105) obtained after enzymatic dissociation of primary and secondary cervicospheres were stained with anti-CD49f-FITC (GoH3; BD Pharmingen), anti-CD71-APC (M-A712; BD Pharmingen), and/or anti-CD133-PE for 60 minutes at 4°C in staining buffer (2% BSA in PBS). Corresponding isotypes were used as control. Cells were washed in PBS, centrifuged, and finally resuspended in 300-μL analysis buffer (1% BSA/2 mmol/L EDTA in PBS). For FACS sorting, 1 × 106 cells were incubated with anti-CD49f-FITC, anti-CD71-APC, and/or anti-CD133-PE, and were finally resuspended in 500-μL analysis buffer. The dead cells and debris were excluded after 7-AAD staining (Invitrogen). Data were collected on FACSAriaIII cell sorter (BD Biosciences) and analyzed using FlowJo software (TreeStar).

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