β actin mab1501

Tissues and cells were homogenized and lysed in lysis buffer as previously described [19 (link)]. The protein concentration was determined by Bradford  protein assay (Bio-Rad). Equal amounts of proteins were loaded and immunoblot analysis was carried out according to standard protocol. The blots were visualized using the ECL chemiluminescence system by Amersham Imager 600 (GE healthcare) and quantified by densitometric analysis using ImageQuantTL (GE healthcare). The following antibodies were used: rabbit polyclonal antibodies against FAM13A (HPA038109; Sigma Life Science); PPARγ (2435), β-Catenin (8480), IRS1 (2382), GSK3β (12456), pGSK3β^S9 (9336), SIRT1 (9475) from Cell Signaling Technology; GAPDH (60004-1-IG, Proteintech); PLIN1 (GP29; Progen Biotechnik GmbH); ATGL (10006409; Cayman Chemical); C/EBPα (sc-61, Santa Cruz); β-ACTIN (MAB1501) from Millipore.

The following primary antibodies were used for this study: Ferritin-H (sc-25617) (lot #J2610 and lot #K0713) from Santa Cruz Biotechnology Inc, USA, α-syn from BD transduction, USA (#610786), β-actin (MAB1501) and RPE65 (MAB5428) from Millipore, USA, LAMP1 (ab24170) and NCOA4 (ab56356) from Abcam, USA, LC3B (#2775), Rab1A (#13075) from Cell Signaling Technology, USA. HRP-conjugated secondary antibodies were purchased from GE healthcare, (NA 931 V, NA 934 V) UK, Alexa 488 and Alexa fluor 546 (A11071, A11018) tagged secondary antibodies used for immunostaining were from Molecular Probes, Invitrogen, USA. cathepsin B assay kit was from immunochemistry, USA and PFD-FDGlu and dextran blue were from Thermo Fisher, USA. Ferric ammonium citrate (FAC) (F5879) and DFO (D9533), bafilomycin A1 (B1793), and other chemicals were from Sigma Aldrich, USA.

Cells were washed twice with phosphate‐buffered saline and harvested in whole‐cell extract lysis buffer (10 mm Tris/HCl pH 7.05, 50 mm NaCl, 1% (w/v) Triton X‐100, 0.5 mΜ PMSF and protease/phospho‐protease inhibitor cocktails by Roche). Protein concentrations were measured with the MicroBCA assay (Thermo Scientific). 40 μg of cell lysates were resolved in 10% (v/v) SDS/PAGE gels and transferred onto nitrocellulose membranes. After blocking, membranes were incubated overnight with primary antibodies. Membranes were then washed and incubated with the appropriate secondary antibody for 90 min at room temperature. Chemiluminescence was detected using Pierce ECL western blotting substrate (Thermo Scientific).
The following antibodies were used in this study: SMAD4 (#9515) by Cell Signaling (Danvers, Massachusetts, USA); TMPRSS6 (ab56180), by Abcam (Cambridge, UK); β‐actin (MAB1501) by Millipore (Burlington, Massachusetts, USA); ferroportin 1 (MTP11) by Alpha Diagnostics (San Antonio, Texas, USA); and STAT3 (sc‐483) and HepC Cag (C7‐50) by Santa Cruz Biotechnology (Dallas, Texas, USA). The polyclonal HCV NS5A and HCV‐1a core antibodies used in this study have been described before [43, 47].
Hepatitis C virus NS5A expression following induction of the cell clones was examined by immunofluorescence with the HCV NS5A polyclonal antibody, as previously described [41].

Total cellular proteins were prepared according to our lab’s protocol [51 (link)]. Tested samples were resolved by SDS-PAGE (10% or 12%) and then underwent immunoblotting. Primary antibodies against cdc2 p34 (sc-54, 1:1000), p-cdc2 p34 (sc-12341, 1:1000), cyclin B1 (sc-594, 1:1000), Cdc25C (sc-327, 1:1000), caspase 3 (sc-56053, 1:500), PARP (sc-7150, 1:2000), Mcl-1 (sc-819, 1:500), and DRP1 (sc-101270, 1:500) were purchased from Santa Cruz Biotechnology (Dallas, TX, USA), Vimentin (10336-1-AP, 1:2000), MMP-2 (10373-2-AP, 1:1000), N-cadherin (22018-1-AP, 1:2000), E-cadherin (20874-1-AP, 1:1000), α-SMA (55135-1-AP, 1:1000), and integrin alpha-4 (19676-1-AP, 1:1000) were purchased from Proteintech (Taipei, Taiwan), and β-actin (MAB1501, 1:10,000) was from Merck Millipore (Burlington, MA, USA).

MET Tyr1234&1235 (#3077), NUMA1 Ser395 (#3429), phospho‐Ser/Thr‐Gln‐Gly (#6966), phospho‐Ser‐Gln (#9607), phospho‐Thr(Asp/Glu)X(Asp/Glu) (#BL4176), phospho‐Thr‐X‐Arg (#2351), CHEK1 Ser345 (#2341), CHEK1 (2G1D5; #2360S), p90RSK Ser380 (#9335), PathScan® (Cell Signaling Technology) Multiplex Western Cocktail I [phospho‐p90RSK, phospho‐Akt, phospho‐p44/42 MAPK (Erk1/2), phospho‐S6; #5301], cleaved caspase‐8 (Asp391; #9496), cleaved caspase‐3 (Asp175; #9661), 53BP1 (#4937), ACIN1 (#4934), cdc2 (#9116T), cell cycle‐dependent kinase 2 (CDK2; #2546T), H2AX (#7631S), 11 (#8967), SMC3 (#5696), and MET (#8198S and #4560) antibodies were all obtained from Cell Signaling Technology (Danvers, MA, USA). Histone H2AX Ser139 (#05‐636), ATM Ser1981 (#05‐740), histone H3 Ser10 (#06‐570), and β‐actin (#MAB1501) antibodies were obtained from Merck Millipore Corporation (Darmstadt, Germany). TIF1B (KAP1) Ser824 (#A300‐767A) antibody was purchased from Bethyl Laboratories, Inc. (Montgomery, TX, USA). SMC3 Ser1083 (#NB100‐653) and ATM (#NB100‐309) antibodies were obtained from Novus Biologicals (Littleton, CO, USA).