Mouse CD117 (c-Kit) eFluor-450

June 20, 2017

Microbiology & Immunology
Columbia University
MD/PhD Student

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Company:

eBioscience™ from Invitrogen Antibodies

Product Name:

CD117 (c-Kit) monoclonal antibody (2B8) eFluor 450

Catalog Number:

48-1171-80

c-Kit (CD117) is an antigen expressed by various populations of hematopoetic stem and progenitor cells. Expression of c-Kit marks key populations, and this antigen is therefore critical for many gating schemes. However, robust detection of c-Kit is often difficult and requires bright fluorophores. We stained mouse bone marrow to analyze expression of transcription factors in progenitor cells. c-Kit efluor 450 was selected as candidate bright FACS antibody that is compatible with complex multi-color panels.

Experimental Design and Results Summary

Applications

Flow Cytometry

Sample

C57BL/6 mouse bone marrow

Primary Incubation

1:100, 30 minutes, 4C

Blocking Agent

FACS buffer (2% FBS in PBS, 2mM EDTA)

Secondary Incubation

N/A

Tertiary Incubation

N/A

Detection

BD Fortessa

Results Summary

Tibia and femurs of C57BL/6 mice were flushed to obtain hematopoetic stem and progenitor cells for FACS analysis of transcription factor expression. Cells were stained with a multi-color FACS panel to discern populations of interest. After gating on live cells, expression of cKit on lineage negative (CD11b, Gr1, Ter119, CD3, CD19, Nk1.1, CD2, CD5-) cells was analyzed. A clear cKit+ population could be detected amongst lineage negative cells. This population could be further analyzed for Sca1 and SLAM expression as per convention stem cell gating schema. Fluorescence intensity of the cKit signal was preserved with aldehyde fixation.

DOI or PMID #

N/A

Additional Notes

N/A

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Summary

The Good

Reasonable separation of cKit+ population

The Bad

Requires countermarker, e.g. lineage, for clearest gating

The Bottom Line

The cKit efluor 450 antibody conjugate provides serviceable detection of cKit expression in mouse bone marrow populations and can be combined into larger panels for stem and progenitor analysis.

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