For the assessment of GSTP1-1 and TRAF2 intracellular levels, a fixed number of synchronized U-2OS cells (∼7.5 × 104 cells) were lysed as previously reported18 (link) and loaded on a 12% SDS-polyacrylamide gel. Proteins were then transferred to PVDF membranes (Millipore, Billerica, MA, USA). For the assessment of LC3-II levels, cell lysates from U-2OS cultures, untreated or treated with CQ (2.5–40.0 μM dose range) for 24 h, were subjected to immunoblot analysis, as previously described.30 (link) A monoclonal anti-GSTP1-1, a polyclonal anti-TRAF2 (Cell Signaling), a polyclonal anti-LC3 (Novus Biologicals, Littleton, CO, USA) and a monoclonal anti-β-actin (Sigma-Aldrich) were used as primary antibodies. Anti-rabbit or anti-mouse secondary antibodies (Cell Signaling) were revealed with the ECL LiteAblot Extend (EuroClone). ImageJ software was used to analyze the band intensities.
Ecl liteablot extend
Manufactured by Euroclone
ECL LiteAblot Extend is a laboratory equipment product. It is designed for chemiluminescence-based detection of proteins in Western blot analyses.
Automatically generated - may contain errors
Lab products found in correlation
Anti rabbit or anti mouse secondary antibodies, by Cell Signaling Technology (2 mentions)
Anti β actin, by Merck Group (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Anti traf2, by Cell Signaling Technology (1 mentions)
Anti lc3, by Novus Biologicals (1 mentions)
Pvdf membrane, by GE Healthcare (1 mentions)
Anti p jnk thr183 tyr185, by Cell Signaling Technology (1 mentions)
Anti jnk, by Cell Signaling Technology (1 mentions)
2 protocols using ecl liteablot extend
1
Western Blot Analysis of Autophagy and Apoptosis Markers
2
Western Blot Analysis of P-JNK
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The cell pellet obtained at each time point was lysed as previously reported
[17 (link)] and the protein concentration
was determined using the Lowry colorimetric assay. Proteins (40 μg) were
loaded on 12% SDS-polyacrylamide gel and transferred onto a PVDF membrane (GE
Healthcare, Chalfont St. Giles, UK). Anti-P-JNK (Thr183 Tyr185) (Cell Signaling,
Beverly, MA, USA), anti-JNK (Cell Signaling) and anti-β actin were used
as primary antibodies. Anti-rabbit or anti-mouse secondary antibodies (Cell
Signaling) were revealed with the ECL LiteAblot Extend (EuroClone). The ImageJ
software was used to analyze the band intensities.
[17 (link)] and the protein concentration
was determined using the Lowry colorimetric assay. Proteins (40 μg) were
loaded on 12% SDS-polyacrylamide gel and transferred onto a PVDF membrane (GE
Healthcare, Chalfont St. Giles, UK). Anti-P-JNK (Thr183 Tyr185) (Cell Signaling,
Beverly, MA, USA), anti-JNK (Cell Signaling) and anti-β actin were used
as primary antibodies. Anti-rabbit or anti-mouse secondary antibodies (Cell
Signaling) were revealed with the ECL LiteAblot Extend (EuroClone). The ImageJ
software was used to analyze the band intensities.
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