Alexa fluor 647 anti xbpis

Cells from the plasma cell assay were collected in a 96-well plate and washed with PBS followed by staining with LIVE/DEAD Fixable Near-IR Dead Cell Stain kit (Invitrogen). Reagent was reconstituted as per manufacturer’s instructions, diluted 1:1000 in PBS, and cells were stained in 100 μL on ice for 20 minutes. The cells were then washed with FACS buffer, PBS (Corning), 2 mM EDTA and 2% heat inactivated fetal bovine serum (Omega Scientific). The cells were then stained with 50 μL antibody cocktail consisting of anti-CD19 (BioLegend), anti-CD45R (BD Pharmingen), and anti-CD138 (BioLegend) in FACS buffer on ice for 15 minutes. Cells were washed twice with 200 μL FACS buffer. XBPIs staining was performed utilizing a Foxp3/transcription factor buffer set (eBioscience) per the manufacturer’s instructions in conjunction with Alexa Fluor 647 anti-XBPIs (BD Biosciences). Cells were resuspended in FACS buffer and combined with an equal volume of 4% paraformaldehyde with 5000 Countbright Absolute Counting Beads (Thermofisher). Stained cells were analyzed on a Fortessa (Becton Dickinson). Division index and data analyses were performed using FlowJo (v10) software (Treestar Inc.).