Pe cy5 anti mouse ly 6a e sca 1 clone d7

The following monoclonal antibodies were employed to perform analytical staining of Lin⁻/Sca-1⁺/c-kit⁺ (SKL cells) and Lin⁻/Sca-1⁺/CD45⁺ (HSCs): fluorescein isothiocyanate (FITC) anti-CD117 (c-kit) (clone2B8; BioLegend, San Diego, CA) and phycoerythrin (PE)-Cy5 anti-mouse Ly-6A/E (Sca-1) (clone D7; eBioscience™, San Diego, CA). All anti-mouse lineage markers (Lin) were conjugated with PE and purchased from BD Biosciences: anti-CD45R/B220 (clone RA3-6B2); anti-Ter-119 (clone TER-119); anti-CD11b (clone M1/70); anti-T-cell receptor β (TCRβ; clone H57-597); anti-Gr-1 (clone RB6-8C5); anti-TCRγδ (clone GL3); and anti-CD45 (clone 30-F11). All monoclonal antibodies were added at saturating concentrations, and the cells were then incubated for 30 min on ice, washed twice, resuspended in RPMI-1640 plus 2% fetal bovine serum, and analyzed with an LSR II flow cytometer (BD, San Diego, CA, USA) [11 (link)].

BMNC staining was performed in medium containing 2 % fetal bovine serum (FBS). All monoclonal antibodies (mAbs) were added at saturating concentrations and the cells were incubated for 30 min on ice, washed twice, resuspended in staining solution at a concentration of 5 × 10⁶ cells/ml, and subjected to analysis using an LSR II (Becton Dickinson, Mountainview, CA). The following anti-mouse Abs were used to detect fluorescein isothiocyanate (FITC)-anti-CD117 (c-Kit) (clone 2B8; BioLegend, San Diego, CA) and Phycoerythrin (PE)-Cy5 anti-mouse Ly-6A/E (Sca-1) (clone D7; eBioscience™, San Diego, CA). All anti-mouse lineage markers (Lin) were conjugated by PE and purchased from BD Biosciences: anti-CD45R/B220 (clone RA3-6B2); anti-Gr-1 (clone RB6-8C5); anti-T-cell receptor β (TCRβ; clone H57-597); anti-TCRγδ (clone GL3); anti-CD11b (clone M1/70); anti-Ter-119 (clone TER-119); and anti-CD34 (clone RAM34) as described [8 –10 (link)].