Total protein was isolated from ESCC tissues and cells using RIPA buffer (Beyotime). The proteins were separated through sodium dodecyl sulfonate‐polyacrylamide gel (SDS‐PAGE; Solarbio) after being quantified via a BCA Protein Quantification Kit (Vazyme). Then, the samples were transferred onto polyvinylidene difluoride membranes (PVDF; Pall Corporation, New York, NYC, USA) and blocked in skim milk for 2 h. Next, the membranes were incubated with primary antibodies against P‐glycoprotein (P‐gp; ab3366; Abcam), glutathione S‐transferase π (GST‐π; ab53942; Abcam), LASP1 (ab156872; Abcam), or GAPDH (ab8245; Abcam) and corresponding secondary antibody (ab150077; Abcam). The protein bands were analyzed with an enhanced chemiluminescence assay kit (Beyotime).
Ab3366
Manufactured by Abcam
Sourced in United States
Ab3366 is an antibody specific for the detection of a target protein. It is designed for use in various laboratory techniques, including immunoassays. The product details and technical specifications are available on request.
Automatically generated - may contain errors
Lab products found in correlation
Ab156872, by Abcam (1 mentions)
Ab53942, by Abcam (1 mentions)
Polyvinylidene fluoride (pvdf), by Pall Corporation (1 mentions)
Ripa buffer, by Beyotime (1 mentions)
Bca protein quantification kit, by Vazyme (1 mentions)
Ab8245, by Abcam (1 mentions)
Ab150077, by Abcam (1 mentions)
Enhanced chemiluminescence assay kit, by Beyotime (1 mentions)
Hrp conjugated goat anti rabbit sa00001 2, by Proteintech (1 mentions)
Chemidoc mp imaging system, by Bio-Rad (1 mentions)
Sc 30621, by Santa Cruz Biotechnology (1 mentions)
Abi 7500 real time pcr system, by Thermo Fisher Scientific (1 mentions)
Mtt assay, by Merck Group (1 mentions)
Verapamil, by Merck Group (1 mentions)
Rhodamine 123, by Merck Group (1 mentions)
4 protocols using ab3366
1
Protein Isolation and Western Blot Analysis
2
Western Blot Immunodetection of Cellular Proteins
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Western blot was performed as described elsewhere [44 (link)]. The primary antibodies that were used were anti-P-glycoprotein (ab3366, RRID: AB_303744), anti-actin (ab11003, RRID: AB_297660) from Abcam (Cambridge, UK), and anti-TRIP6 (HPA052813, RRID: AB_2681961) from Atlas Antibodies (Bromma, Sweden). SuperSignal™ West Pico PLUS Chemiluminescent Substrate or West Femto Maximum Sensitivity Substrate (Pierce, Rockford, IL, USA) were applied on a membrane to detect the chemiluminescence signal of secondary HRP-conjugated goat anti-mouse (SA00001-1, RRID: AB_272565) and HRP-conjugated goat anti-rabbit (SA00001-2, RRID: AB_272564) from Proteintech (Rosemont, IL, USA). Images were obtained using ChemiDoc MP imaging system (Biorad, Hercules, CA, USA).
3
Quantitative Analysis of Hepatic Genes
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Total RNA was prepared using TRIzol reagent. SYBR Green-based qRT-PCR was performed with an ABI 7500 Real-Time PCR System (Applied Biosystems Inc., Hercules, CA, USA). Data were normalized against a cyclophilin control. The qRT-PCR primer sequences are presented in Supplementary Table 1 .
Specific antibodies against HBx (MAB8419; Millipore, Danver, MA, USA), PXR (PP-H4417-00; R&D Systems, Minneapolis, MN, USA), MDR1 (ab3366; Abcam, Cambridge, UK), CYP3A4 (H00001576-B01P; Novus Biologicals, Littleton, CO, USA), SULT2A1 (ab38416; Abcam, Cambridge, UK), CYP3A (sc-30621; Santa Cruz Biotechnology, Dallas, TX, USA), cytochrome p450 1A2 (CYP1A2) (AP11325c; Abgent, Suzhou, China), and GSTM1 (AP6896b; Abgent, Suzhou, China) were used.
Specific antibodies against HBx (MAB8419; Millipore, Danver, MA, USA), PXR (PP-H4417-00; R&D Systems, Minneapolis, MN, USA), MDR1 (ab3366; Abcam, Cambridge, UK), CYP3A4 (H00001576-B01P; Novus Biologicals, Littleton, CO, USA), SULT2A1 (ab38416; Abcam, Cambridge, UK), CYP3A (sc-30621; Santa Cruz Biotechnology, Dallas, TX, USA), cytochrome p450 1A2 (CYP1A2) (AP11325c; Abgent, Suzhou, China), and GSTM1 (AP6896b; Abgent, Suzhou, China) were used.
4
Evaluating Multidrug Resistance Pathways
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DOX was obtained from Pfizer Italia Srl (Rome, Italy). Geniposide was obtained from the National Institute for the Control of Pharmacological and Biological Products (Beijing, China). Rhodamine 123 (Rho123), MTT assay and verapamil were purchased from Sigma-Aldrich, Merck Millipore (Darmstadt, Germany). Monoclonal antibodies against ABCB1 (ab3366) were purchased from Abcam, Ltd., (Hong Kong, China).
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