Isolation and Characterization of Mouse Muscle Stem Cells

Mammary tumor-bearing PyMT+ and Neu+ female mice, and age-matched non-tumor-bearing control females were sacrificed with CO₂. Approximately 0.5 grams of muscles were dissected from the hind limb of each mouse. Mouse muscle dissociation kit (130–098-305) and mouse satellite cell isolation kit (130–104-268) were used to isolate mouse muscle stem cells (MuSC) following MACS Miltenyi Biotec manufacture’s protocols. Purified MuSC were stained with antibodies CD54-APC (116120; Biolegend) and CD82-PE (PA5–13228; Invitrogen) and analyzed as described previously ^{26 (link)}. Stained cells were acquired using a BD LSR II flow cytometer and data were analyzed using FlowJo software. Forward and side scatter were used to ensure that only live cells were considered in the analysis. Gating was done using appropriate PE (555749) and APC (555576) (BD Pharmingen) isotype control antibodies.

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Wang R., Kumar B., Bhat-Nakshatri P., Prasad M.S., Jacobsen M.H., Ovalle G., Maguire C., Sandusky G., Trivedi T., Mohammad K.S., Guise T., Penthala N.R., Crooks P.A., Liu J., Zimmers T, & Nakshatri H. (2020). Aging-associated skeletal muscle defects in HER2/Neu transgenic mammary tumor model. JCSM rapid communications, 4(1), 24-39.

Publication 2020

CD54-APC BD LSR II flow cytometer

Antibodies Cd54 Cells Females Hind limb Isolation Isotype Macs protocols Mammary tumor Mouse Muscle Muscle stem cells Tumor

Corresponding Organization : Richard L. Roudebush VA Medical Center

Other organizations : University of Arkansas for Medical Sciences

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Variable analysis

independent variables

Mammary tumor-bearing PyMT+ and Neu+ female mice
Age-matched non-tumor-bearing control females

dependent variables

Purified mouse muscle stem cells (MuSC) characterized by expression of CD54 and CD82

control variables

Approximately 0.5 grams of muscles dissected from the hind limb of each mouse
Mouse muscle dissociation kit (130–098-305) and mouse satellite cell isolation kit (130–104-268) used to isolate MuSC
Appropriate PE (555749) and APC (555576) isotype control antibodies used for gating

controls

Positive control: Mammary tumor-bearing PyMT+ and Neu+ female mice
Negative control: Age-matched non-tumor-bearing control females

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