Whole cell lysates were obtained using the M-PER Mammalian Protein Extraction Reagent (Pierce) plus several protease inhibitors (1 mM PMSF; 10 μg/ml leupeptin, 2 μg/ml pepstatin). Cell lysates were then loaded (40 μg/lane) in a 4–12% SDS–PAGE gel to separate proteins and transferred to nitrocellulose membrane. Antibodies to SIRT1 (H-300, 1:500) and β-actin (Sigma–Aldrich, 1:1000) were used for detection. The optical density of the SIRT1 and β-actin bands was quantified using the ImageJ software. The values are presented as the ratio of SIRT1 and Actin optical density. Data are representative of three independent experiments.
H 300
Manufactured by Merck Group
The H-300 is a laboratory equipment product designed for general use in research and analytical applications. It serves as a compact and versatile unit capable of performing various functions essential to laboratory workflows.
Automatically generated - may contain errors
Lab products found in correlation
β actin, by Merck Group (1 mentions)
Ab5821, by Abcam (1 mentions)
Fluorescein isothiocyanate (fitc), by Jackson ImmunoResearch (1 mentions)
Chk2 pthr 68, by Cell Signaling Technology (1 mentions)
Atm pser 1981, by Cell Signaling Technology (1 mentions)
Img 124a, by Novus Biologicals (1 mentions)
Bu 33, by Merck Group (1 mentions)
Cyanine3 (cy3), by Jackson ImmunoResearch (1 mentions)
2 protocols using h 300
1
Immunoblotting for SIRT1 Expression
2
Immunofluorescence Analysis of Cellular Markers
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Antibodies included fibrillarin 1∶1000 (Abcam ab18380 or ab5821), CENP-A 1∶500 (Abcam 13939 or Upstate 07-574), TRF2 1∶200 (Imgenex IMG-124A or Novus NB110-57130), Ki67 antigen 1∶500 (Novocastra Laboratories Ltd.), H2AX-p 1∶300 (Millipore 05-636 or Abcam ab2893), ATM-p Ser1981 1∶250 (Cell Signaling 5883), Chk2-p Thr68 1∶250 (Cell Signaling 2661), SMC2 1∶300 (Cell Signaling 5394), SMC4 1∶300 (Cell Signaling 5547), goat polyclonal to DDDDK (FLAG) tag 1∶500 (Abcam ab1257), UBF 1∶150 (Santa Cruz H-300), and anti-BrdU 1∶400 (Sigma clone BU-33). Primary antibodies were detected using anti-mouse, anti-rabbit, or anti-goat secondary antibodies conjugated to Alexa Fluor 488, 594, 647 (Molecular Probes), FITC, Cy3, or Cy5 (Jackson Immunoresearch, Inc.).
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