Anti cd49f apc

The dorsal skin was dissected and laid flat for subcutaneous fat
scratching and then floated epidermis side up in 0.25 % Trypsin (Gibco) in
PBS for 1 hour at 37°C to separate epidermal and dermal layers.
Following the incubation, the epidermis was carefully scraped off and
transferred to 10% FBS (fetal bovine serum) in DMEM (Gibco), in which it was
manually gently dissociated using a serological pipette. The entire volume
was filtered through a 70 μm cell strainer (VWR) to obtain a single
cell suspension that was then centrifuged and resuspended in 2 % FBS in PBS.
For cell staining, antibodies were directly added to the cell suspension and
incubated for 10 min on ice. Afterwards, cells were washed using 2 % FBS PBS
and resuspended for sorting in 5 mM EDTA PBS. The following antibodies were
used: anti-CD49f-APC (1:300, 313615, BioLegend), anti-CD49f-PercP/Cy5.5
(1:300, 313618, BioLegend), anti-CD49f-FITC (1:300, 313606, BioLegend),
anti-CD34-PE (1:50, 551387, BD Pharmingen), anti-CD34-Alexa Fluor 647 (1:50,
560230, BD Pharmingen), anti-Sca1-Ly-6A/E-Violet 605 (1:200, 108133,
BioLegend) and DAPI (1:1000, Biotium). Flow cytometry was performed on a BD
LSRII cytometer (BD Biosciences), sortings on a BD FACS Aria II sorter (BD
Biosciences). Flow cytometry data was collected and exported using BD FACs
Diva software (BD Biosciences) and analyzed and plotted using FlowJo
software.