Cells were fixed in ice-cold 70% ethanol, and incubated overnight at 4 °C. Subsequently, cells were stained for rabbit anti-phospho-histone H3 antibody (1:100, Cell Signalling, #9701) and mouse MPM2 (1:100, Millipore, 05-368). After washing with PBS–0.05% Tween-20, cells were incubated with Alexa 488-conjugated and Alexa 647-conjugated secondary antibodies (1:100, Molecular Probes) and counterstained with propidium iodide/RNase (Sigma). FACS analyses were performed on a FACS-Calibur (Becton Dickinson) and samples were analyzed using Cell Quest software. Data were analyzed using FlowJo software. At least 10,000 events were analyzed per sample.
Rabbit anti phospho histone h3 antibody
Manufactured by Cell Signaling Technology
Rabbit anti-phospho-histone H3 antibody is a primary antibody that specifically recognizes histone H3 phosphorylated at serine 10 or serine 28. It is commonly used in immunohistochemistry and Western blotting applications to detect and quantify mitotic cells.
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2 protocols using rabbit anti phospho histone h3 antibody
1
Cell Cycle Analysis by Flow Cytometry
2
Insect Gut Immunostaining Protocol
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The intestines were dissected in Insect Medium. Gut fixation and immunostraining were as previously described (Linetal.,2008). Following antisera and dyes were used: rabbit anti-phospho-Histone H3 antibody (Cell signaling, 1:1000); mouse anti-GFP antibody (Invitrogen, 1:1000); Alexa-488-conjugated goat anti-mouse secondary antibodies and Alexa-568-conjugated goat anti-rabbit secondary antibodies (Invitrogen, 1:1000); DAPI (49,69-diamidino-2-phenylindole, Sigma; 0.1 mg/ml, 5 min incubation). Images were captured by confocal microscopy (a Leica SP5 confocal microscope). All images were processed in Adobe Photoshop.
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