Biotin Rat Anti-Mouse CD106

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Propidium iodide (PI) (1 μg/ml, Sigma #P4170) was added to differentiate between live and dead cells. Only live cells (PI−) were counted.	FACS analysis and cell sorting were performed in a BD FACSAriaII (BD Biosciences, San Diego, CA) using the FACSDiva software (BD Biosciences). Single-cell precision was used for sorting single cells into 96-well plate for clonal analysis, and 4-way purity precision was used for bulk sort.

Protocol tips
Propidium iodide (PI) (1 μg/ml, Sigma #P4170) was added to differentiate between live and dead cells. Only live cells (PI−) were counted.
Downstream tips
FACS analysis and cell sorting were performed in a BD FACSAriaII (BD Biosciences, San Diego, CA) using the FACSDiva software (BD Biosciences). Single-cell precision was used for sorting single cells into 96-well plate for clonal analysis, and 4-way purity precision was used for bulk sort.

Publication protocol

Dissociated cells were incubated with antibodies (APC anti-α7-Integrin, AbLab #67-0010-05, Vancouver, Canada; APC anti-β1-Integrin, eBioscience #17-0291-82; RRID:AB_1210793, San Diego, CA; PE-Cy7 anti-CD31, BD Biosciences Cat#561410; RRID:AB_10612003, San Jose, CA; Biotin anti-CD34, eBioscience Cat#13-0341-81; RRID:AB_466424; PE-Cy7 anti-CD45, BD Biosciences Cat#552848; RRID:AB_394489; Biotin anti-CXCR4, eBioscience Cat#13-9991-82; RRID:AB_10609202; Biotin anti-VCAM-1, BD Biosciences Cat#553331; RRID:AB_10053328; PE streptavidin, BD Biosciences Cat#554061; all at 0.5 μl per 1 million cells) on ice for 30 min. Propidium iodide (PI) (1 μg/ml, Sigma #P4170) was added to differentiate between live and dead cells. Only live cells (PI−) were counted. FACS analysis and cell sorting were performed in a BD FACSAriaII (BD Biosciences, San Diego, CA) using the FACSDiva software (BD Biosciences). Single-cell precision was used for sorting single cells into 96-well plate for clonal analysis, and 4-way purity precision was used for bulk sort. Data were analyzed using FlowJo (FLOWJO LLC, Ashland, OR). Further information on antibodies used is listed in Key Resources Table.

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Manufacturer protocol

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