The cells were stimulated for 5 h with anti-CD3/28 beads (Miltenyi Biotec, Auburn, CA, USA) and stained according to the manufacturer’s instructions. Cells were analyzed using an LSRFortessa cell analyzer (BD, Mississauga, ON, Canada). The gating strategy is displayed in Supplementary Figure 1
Anti cd27 apc vio770
Manufactured by Miltenyi Biotec
Sourced in United States
Anti-CD27-APC-Vio770 is a fluorochrome-conjugated antibody that binds to the CD27 cell surface marker. CD27 is a co-stimulatory receptor expressed on T cells and B cells.
Automatically generated - may contain errors
Lab products found in correlation
Anti cd3 28 beads, by Miltenyi Biotec (1 mentions)
Anti cd38 fitc, by Miltenyi Biotec (1 mentions)
Vx software, by FlowJo (1 mentions)
Anti pd 1 fitc, by BioLegend (1 mentions)
Anti ccr7 apc, by BioLegend (1 mentions)
Lsrfortessa cell analyzer, by BD (1 mentions)
Anti cd138 pe, by Miltenyi Biotec (1 mentions)
Anti cd16 fitc, by Miltenyi Biotec (1 mentions)
Anti cd3 fitc, by Miltenyi Biotec (1 mentions)
Anti cd14 fitc, by Miltenyi Biotec (1 mentions)
2 protocols using anti cd27 apc vio770
1
Quantification of Circulating CXCR5+ T Cells
2
Isolation of Immune Cell Subsets from CSF
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The index patients’ parents have given written informed consent, and analyses were approved by the Charité University Hospital Institutional Review Board.
4 ml of CSF was collected during the acute phase of encephalitis (Nikolaus et al., 2018 (link)) and immediately processed for cell pellet cryopreservation, therefore centrifuged for 10 min at 400× g, with supernatant stored at −80°C and the pellet resuspended in 500 µl of 10% dimethyl sulfoxide, 45% fetal calf serum, 45% RPMI medium before freezing at −80°C. PPE was collected 6 d after lumbar puncture and stored at −80°C.
We used fluorescence-activated cell sorting to isolate single CD138+ ASCs, CD20+CD27+ MBCs, and CD20+CD27− NMBCs from preselected viable CD3−CD14−CD16−DAPI− lymphocytes into 96-well PCR plates. The following antibodies were applied: anti–CD3-FITC (1:25; Miltenyi Biotec; #130-098-162), anti–CD14-FITC (1:25; Miltenyi Biotec; #130-098-063), anti–CD16-FITC (1:25; Miltenyi Biotec; #130-098-099), anti–CD20-PerCP-Vio700 (1:50; Miltenyi Biotec; #130-100-435), anti–CD27-APC-Vio770 (1:12.5; Miltenyi Biotec; #130-098-605), and anti–CD138-PE (1:50; Miltenyi; #130-098-122).
4 ml of CSF was collected during the acute phase of encephalitis (Nikolaus et al., 2018 (link)) and immediately processed for cell pellet cryopreservation, therefore centrifuged for 10 min at 400× g, with supernatant stored at −80°C and the pellet resuspended in 500 µl of 10% dimethyl sulfoxide, 45% fetal calf serum, 45% RPMI medium before freezing at −80°C. PPE was collected 6 d after lumbar puncture and stored at −80°C.
We used fluorescence-activated cell sorting to isolate single CD138+ ASCs, CD20+CD27+ MBCs, and CD20+CD27− NMBCs from preselected viable CD3−CD14−CD16−DAPI− lymphocytes into 96-well PCR plates. The following antibodies were applied: anti–CD3-FITC (1:25; Miltenyi Biotec; #130-098-162), anti–CD14-FITC (1:25; Miltenyi Biotec; #130-098-063), anti–CD16-FITC (1:25; Miltenyi Biotec; #130-098-099), anti–CD20-PerCP-Vio700 (1:50; Miltenyi Biotec; #130-100-435), anti–CD27-APC-Vio770 (1:12.5; Miltenyi Biotec; #130-098-605), and anti–CD138-PE (1:50; Miltenyi; #130-098-122).
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