Flow cytometry Anti-bodies Human - CD70

Flow cytometry is an immunophenotyping technique whereby sing cell suspensions are stained for either cell surface markers or intracellular proteins by fluorescently-labelled antibodies and analyzed with a flow cytometer, where fluorescently-labelled molecules are excited by the laser to emit light at varying wavelengths, which is then detected by the instrument. There are several key criteria which are required to be kept in mind while designing a flow experiment- 1. Antibody titration (optimal dilution of antibodies should be calculated in order to avoid over- or under- saturated signals for proper detection of surface and intracellular markers), 2. Precision (3 or more replicates of the sample should be used per experiment), 3. Specificity (proper isotype controls should be included in the experiment), 4. Day-to-day variability (experiments should be repeated 3 or more times to ensure consistency and avoid variability due to flow cytometer settings), 5. Antibody interaction (Fluorescence minus one or FMO should be used, which is the comparison of signals from panel minus one antibody vs. the full panel), and 6. Antibody stability (fluorescently-labelled antibodies should be stored at 4C).

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Found 3 matching solutions for this experiment

FITC Mouse Anti-Human CD70

BD Biosciences

Upstream tips	Protocol tips	Downstream tips
	PBMC from patients with autoimmune diseases or from cultures of 5-azaC-treated PBMC from healthy donors were incubated in phosphate-buffered saline (PBS)/0.001% azide containing 10% horse serum at 4°C for 30 min to block non-specific binding. All staining procedures were performed at 4°C in the dark. The cells were then washed, incubated for 60 min, with a single or mixture of Ab.	Fluorescence-activated cell sorting (FACS) analyses were performed using a FACS ARIA IIIU flow cytometer and FACSDiva software V.6.1.3 (Becton Dickinson, New Jersey, USA) or an iCyte Synergy (Sony Biotechnology, San Jose California, USA) and WINLIST V.8 (Verty Software House, http://www.vsh.com) software.

Protocol tips
PBMC from patients with autoimmune diseases or from cultures of 5-azaC-treated PBMC from healthy donors were incubated in phosphate-buffered saline (PBS)/0.001% azide containing 10% horse serum at 4°C for 30 min to block non-specific binding. All staining procedures were performed at 4°C in the dark. The cells were then washed, incubated for 60 min, with a single or mixture of Ab.
Downstream tips
Fluorescence-activated cell sorting (FACS) analyses were performed using a FACS ARIA IIIU flow cytometer and FACSDiva software V.6.1.3 (Becton Dickinson, New Jersey, USA) or an iCyte Synergy (Sony Biotechnology, San Jose California, USA) and WINLIST V.8 (Verty Software House, http://www.vsh.com) software.

Manufacturer protocol Publication protocol 1 Relevant paper

PE Mouse Anti-Human CD70

BD Biosciences

Upstream tips	Protocol tips	Downstream tips
	Adherent cells were washed with PBS and incubated in Cell Dissociation Buffer (Thermo Fisher Scientific) for 20 min in a 37°C incubator to detach cells. After centrifugation, the cells were collected and counted. Cells were incubated either with or without Ab for 1 h on ice and then cells were washed three times in PBS.	Cells were analyzed with an LSRFortessa flow cytometer (BD, Heidelberg, Germany) and FlowJo software (FlowJo, Ashland, OR, USA).

Protocol tips
Adherent cells were washed with PBS and incubated in Cell Dissociation Buffer (Thermo Fisher Scientific) for 20 min in a 37°C incubator to detach cells. After centrifugation, the cells were collected and counted. Cells were incubated either with or without Ab for 1 h on ice and then cells were washed three times in PBS.
Downstream tips
Cells were analyzed with an LSRFortessa flow cytometer (BD, Heidelberg, Germany) and FlowJo software (FlowJo, Ashland, OR, USA).

Manufacturer protocol Publication protocol 1 Relevant paper

CD70 Monoclonal Antibody (BU69), FITC

eBioscience

Upstream tips	Protocol tips	Downstream tips
	cells (2×106) along with blank controls were firstly centrifuged at 2 000 rpm for 10 min, rinsed in PBS, and Ab added. After mixture and washing, cells were fixed by 0.5 mL paraformaldehyde and loaded on flow cytometry for detection.

Protocol tips
cells (2×106) along with blank controls were firstly centrifuged at 2 000 rpm for 10 min, rinsed in PBS, and Ab added. After mixture and washing, cells were fixed by 0.5 mL paraformaldehyde and loaded on flow cytometry for detection.

Manufacturer protocol Publication protocol 1 Relevant paper

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