Isolating Adipose-Derived Stem Cells from Murine Periaortic Adipose Tissue

Abstract

Source: Qi, Y., et al. Isolation, Culture, and Adipogenic Induction of Neural Crest Original Adipose-Derived Stem Cells from Periaortic Adipose Tissue. J. Vis. Exp. (2020)

This video demonstrates a technique for isolating and culturing fluorescently labeled neural crest-derived adipose-derived stem cells (ADSCs) from murine periaortic arch adipose tissue. Following isolation, the NCADSCs are detected, sorted using fluorescence-activated cell sorting, and subsequently cultured on a plate, promoting cell proliferation.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines.

1. Isolation of the stromal vascular fraction (SVF)

Collect the periaortic arch adipose tissue (PAAT) of 5-6 mice into one 2 mL microcentrifuge tube containing 1 mL freshly prepared digestion medium and mince the tissue using surgical scissors in an Eppendorf tube at room temperature (RT).

1. Transfer the mix into 50 mL tubes containing 9 mL of the digestion medium. Homogenize the tissues by pipetting up and down with a 1 mL pipette 10x.
2. Incubate the tubes at 37
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Results

Figure 1: Adipogenic differentiation of NCADSCs isolated from PAAT. (A) General gating scheme for characterizing and sorting NCADSCs (RFP) populations. (B) Fluorescence microscope images show that the NCADSCs adhere.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
BSA	VWR life sciences	0332-100G	50 mg/mL working concentration; lot #: 0536C008
Collagenase, Type I	Gibco	17018029
Erythrocyte Lysis Buffer	Invitrogen	00-4333
FBS	Corning	R35-076-CV	50 mg/mL working concentration; lot #: R2040212FBS
HBSS	Gibco	14025092
HDMEM	Gelifesciences	SH30243.01	Lot #: AD20813268
PBS (Phosphate buffered saline)	ABCONE (China)	P41970
Penicillin-Streptomycin	Gibco	15140122