Mitochondrial-specific assessment of superoxide was performed as has been previously described (Case et al., 2016 (link)). Briefly, cells were stained with cell-type specific fluorescent antibodies [anti-CD3ε PE-Cy7 (clone 145–2C11, eBioscience), CD19 APC-Cy7 (clone 6D5, BioLegend), CD11b SB-436 (clone M1/70, eBioscience), CD11c APC (clone N418, eBioscience), and NK1.1 SB-600 (clone PK136, eBioscience)] in addition to 1 µM of (MATH) -sensitive mitochondrial-localized probe, MitoSOX Red (#M36008, Thermo Fisher Scientific) for 30 min at 37 °C. Cells were analyzed on an LSRII flow cytometer at 488/610 nm excitation/emission, respectively, and data analyzed using FlowJo software.
Cd19 apc cy7 clone 6d5
Manufactured by BioLegend
CD19 APC-Cy7 (clone 6D5) is a fluorescently labeled antibody that binds to the CD19 surface marker. CD19 is a transmembrane protein expressed on B cells and can be used to identify and enumerate B cell populations.
Automatically generated - may contain errors
Lab products found in correlation
Cd11c apc clone n418, by Thermo Fisher Scientific (2 mentions)
Cd11b sb 436 clone m1 70, by Thermo Fisher Scientific (2 mentions)
Nk1.1 sb 600 clone pk136, by Thermo Fisher Scientific (2 mentions)
Mitosox red, by Thermo Fisher Scientific (1 mentions)
Mitosox red mitochondrial superoxide indicator, by Thermo Fisher Scientific (1 mentions)
Cd3ε pe cy7, by BioLegend (1 mentions)
Lsr 2 flow cytometer, by BD (1 mentions)
2 protocols using cd19 apc cy7 clone 6d5
1
Mitochondrial Superoxide Assessment in Cells
2
Splenocyte Mitochondrial Superoxide Assay
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Freshly isolated live splenocytes were incubated in RPMI supplemented with the following fluorescently tagged antibodies targeting extracellular proteins: CD3ε PE-Cy7, CD19 APC-Cy7 (clone 6D5; BioLegend), CD11b SB-436 (clone M1/70; eBioscience), CD11c APC (clone N418; eBioscience), and NK1.1 SB-600 (clone PK136; eBioscience). Concurrently, 1 μM MitoSOX Red mitochondrial superoxide indicator (Thermo Fisher Scientific) was added, and cells were incubated for 30 minutes at 37 °C. Cells were washed and resuspended in phosphate-buffered saline, and data were acquired using a customized BD LSR II flow cytometer. MitoSOX Red mean fluorescence intensities were normalized to intraexperiment sham-operated control samples. All flow cytometry experiments were conducted with accompanying single-color and fluorescence-minus-one control tubes. Data were analyzed using FlowJo software.
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