Immunoblotting, immunofluorescence and FACS assays were performed as previously described (14 (link)). Antibodies that were employed to perform these studies were the following: p53 (1:500; mouse monoclonal; cat. no. PIMA512557; Invitrogen/Thermo Fisher Scientific, Waltham, MA, USA), p21 (1:500; mouse monoclonal; cat. no. AHZ0422; Thermo Fisher Scientific), centrin 20H5 (1:1,000; mouse monoclonal; Mayo Clinic, Rochester, MN, USA), pericentrin (1:500; rabbit polyclonal; cat. no. ab4448l) and p-retinoblastoma (1:400; rabbit polyclonal; cat. no. ab47763; both from Abcam, Cambridge, MA, USA), γ-tubulin (1:4,000; mouse monoclonal; cat. no. MA1-850; Thermo Fisher Scientific), Aurora-A (1:500; mouse monoclonal; cat. no. ab13824; Abcam), p-Aurora-A (1:400; rabbit monoclonal; cat. no. 3079S; Cell Signaling Technology, Inc., Danvers, MA, USA), p-retinoblastoma (1:500; mouse monoclonal; cat. no. R6878-1ML) and β-actin (1:5,000; mouse monoclonal, cat. no. A2228-100UL; both from Sigma-Aldrich, St. Louis, MO, USA). Results were derived from three independent experiments with comparable outcomes.
Protocol detail
Immunoblotting, Immunofluorescence, and FACS Protocols
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Actin
Antibodies
Assays
Aurora a
Centrin
Immunofluorescence
Mouse
Pericentrin
Rabbit
Retinoblastoma
Tubulin
Corresponding Organization :
Other organizations : Mayo Clinic, Beth Israel Deaconess Medical Center, Harvard University
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Variable analysis
independent variables
- P53 (1:500; mouse monoclonal; cat. no. PIMA512557; Invitrogen/Thermo Fisher Scientific, Waltham, MA, USA)
- P21 (1:500; mouse monoclonal; cat. no. AHZ0422; Thermo Fisher Scientific)
- Centrin 20H5 (1:1,000; mouse monoclonal; Mayo Clinic, Rochester, MN, USA)
- Pericentrin (1:500; rabbit polyclonal; cat. no. ab4448l)
- P-retinoblastoma (1:400; rabbit polyclonal; cat. no. ab47763; both from Abcam, Cambridge, MA, USA)
- γ-tubulin (1:4,000; mouse monoclonal; cat. no. MA1-850; Thermo Fisher Scientific)
- Aurora-A (1:500; mouse monoclonal; cat. no. ab13824; Abcam)
- P-Aurora-A (1:400; rabbit monoclonal; cat. no. 3079S; Cell Signaling Technology, Inc., Danvers, MA, USA)
- P-retinoblastoma (1:500; mouse monoclonal; cat. no. R6878-1ML)
- β-actin (1:5,000; mouse monoclonal, cat. no. A2228-100UL; both from Sigma-Aldrich, St. Louis, MO, USA)
- Immunoblotting, immunofluorescence and FACS assays
- Immunoblotting, immunofluorescence and FACS assays were performed as previously described (14 (link)).
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