Total proteins from the cell extracts were separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, Bedford, MA, USA). After probing with primary antibodies, membranes were washed in Tris-buffered saline containing 0.05% Tween-20 (TBST) and incubated with a horse-radish peroxidase-linked secondary antibody. Finally, Western signals were detected using HRP substrate luminal reagent (Millipore). The primary antibodies used are provided in the supplemental methods.
Hrp substrate luminal reagent
Manufactured by Merck Group
The HRP substrate luminal reagent is a laboratory product designed to detect the presence of horseradish peroxidase (HRP) in various assays. It serves as a substrate for the HRP enzyme, emitting chemiluminescence upon reaction, which can be measured to quantify the amount of HRP present in the sample.
Automatically generated - may contain errors
3 protocols using hrp substrate luminal reagent
1
Protein Separation and Western Blot Analysis
2
Protein Separation and Western Blot Analysis
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Total proteins from the cell extracts were separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, Bedford, MA, USA). After probing with primary antibodies, membranes were washed in Tris-buffered saline containing 0.05% Tween-20 (TBST) and incubated with a horse-radish peroxidase-linked secondary antibody. Finally, Western signals were detected using HRP substrate luminal reagent (Millipore). The primary antibodies used are provided in the supplemental methods.
3
Protein Sample Analysis by Western Blot
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Equal amount of protein samples were resolved in SDS-PAGE. Antibody to BAP1 (ab199396) was obtained from Abcam, Cambridge, Massachusetts; Antibody to Flag (F1804) was obtained from Sigma-Aldrich St. Louis, MO, Missouri; Antibodies to Fibronectin (sc-8422), c-Met (sc-8057) and GAPDH (sc-25778) were from Santa Cruz Biotechnology, Santa Cruz, California; those to ubiquityl-histone H2A (Lys119) (#8240) and beta-actin (#4970) were from Cell Signaling, Boston, Massachusetts. Western blot signals were measured using HRP substrate luminal reagent (Millipore, Billerica, Massachusetts).
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