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2 protocols using anti prb807 811

1

Immunostaining of Cardiac Muscle Cells

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Staining was performed as described28 (link),55 (link),56 (link). Primary antibodies: mouse monoclonal anti-mCherry (1:200, Clontech, 632543 or 1:50, DSHB, 3A11), anti-Tropomyosin (1:200, Sigma, T9283), anti-sarcomeric alpha Actinin (1:100, Abcam, ab9465), anti-Aurora B (1:200, BD Transduction Laboratories, 611083), anti-p27 (1:50, BD Transduction Laboratories, 610242), anti-Ki67 (1:250, Abcam, ab8191), rabbit polyclonal anti-Troponin I (sc-15368), anti-Cyclin A (sc-751), anti-cdc2 (sc-954), anti-Geminin (sc-13015) (all 1:50, Santa Cruz), anti-phospho-Histone H3 (Ser10) (1:200, Millipore, 06-570), anti-pRb807/811 (1:100, Cell Signaling, 9308), anti-mAG (1:300, MBL, PM011), anti p27 (1:50, SantaCruz, sc-528), anti-survivin (1:50, Novus, NB500-201K8), rat monoclonal anti-BrdU (1:100, Abcam, ab6326) and goat polyclonal anti c-kit (1:100, R&D Systems, AF1356). Immune complexes were detected with ALEXA 488- or ALEXA 594-conjugated secondary antibodies (1:200; Molecular Probes). DNA was visualized with DAPI (4′, 6′-diamidino-2-phenylindole, 0.5 g/ml). For BrdU, cells were cultured in 30 μM BrdU (neonatal: last 24 h, adult: last 5 days).
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2

Multiparameter Immunofluorescence Staining

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Staining was performed as described (24 (link)). Primary antibodies: anti-tropomyosin (1:200, Sigma), anti-actinin (1:100, Abcam), anti-aurora B (1:200) (both BD Transduction Laboratories), rabbit polyclonal anti-troponin I, anti-cyclin A, anti-cyclin dependent kinase 1 (cdc2), anti-geminin (all 1:50, Santa Cruz Biotechnology), anti-phospho-histone H3 (Ser10) (1:200, Millipore), anti-pRb807/811 (1:100, Cell Signaling), anti-mAG (1:300, MBL), rat monoclonal anti-5-Bromo-2-deoxyuridine (BrdU) (1:100, Abcam). Immune complexes were detected with ALEXA 488- or ALEXA 594-conjugated secondary antibodies (1:200; Molecular Probes). DNA was visualized with DAPI'(4” 6'-diamidino-2-phenylindole, 0.5 μg/ml). For BrdU, cells were cultured in 30 μM BrdU (Sigma) (neonatal: last 24–48 h, adult: last 5 days).
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