Purified His6‐Larg1 was used to raise mouse‐specific antibodies using a standard protocol (Pocono Rabbit Farm & Laboratory). The antibodies were affinity‐purified as previously described
65 (link). For immunoblotting, samples resolved by SDS‐PAGE were transferred onto 0.2 μm nitrocellulose membranes (cat# 1620112; Bio‐Rad), which were blocked with 5% nonfat milk at room temperature for 1 h before being incubated with the appropriate primary antibodies: anti‐Flag (cat# F1804; Sigma‐Aldrich), 1: 5000; anti‐HA (cat# H3663; Sigma‐Aldrich), 1:5000; anti‐tubulin (E7; DSHB) 1:10,000; anti‐ADPR (cat# MABE1016; Sigma‐Aldrich), 1:1000; anti‐ICDH
66 (link), 1:10,000; anti‐PDHA1 (cat# 18068‐1‐AP; Proteintech), 1:5000; anti‐ATPB (cat# 17247‐1‐AP; Proteintech), 1:2000; anti‐TOM20 (cat# 11802‐1‐AP; Proteintech), 1:5000; anti‐VDAC1 (cat# 55259‐1‐AP; Proteintech), 1:2000; anti‐Cyto‐c (cat# sc‐13560; Santa Cruz), 1:1000; anti‐Larg1, 1:2500. Membranes were then incubated with appropriate IRDye infrared secondary antibodies and scanned by an Odyssey infrared imaging system (Li‐Cor's Biosciences).
65 (link). For immunoblotting, samples resolved by SDS‐PAGE were transferred onto 0.2 μm nitrocellulose membranes (cat# 1620112; Bio‐Rad), which were blocked with 5% nonfat milk at room temperature for 1 h before being incubated with the appropriate primary antibodies: anti‐Flag (cat# F1804; Sigma‐Aldrich), 1: 5000; anti‐HA (cat# H3663; Sigma‐Aldrich), 1:5000; anti‐tubulin (E7; DSHB) 1:10,000; anti‐ADPR (cat# MABE1016; Sigma‐Aldrich), 1:1000; anti‐ICDH
66 (link), 1:10,000; anti‐PDHA1 (cat# 18068‐1‐AP; Proteintech), 1:5000; anti‐ATPB (cat# 17247‐1‐AP; Proteintech), 1:2000; anti‐TOM20 (cat# 11802‐1‐AP; Proteintech), 1:5000; anti‐VDAC1 (cat# 55259‐1‐AP; Proteintech), 1:2000; anti‐Cyto‐c (cat# sc‐13560; Santa Cruz), 1:1000; anti‐Larg1, 1:2500. Membranes were then incubated with appropriate IRDye infrared secondary antibodies and scanned by an Odyssey infrared imaging system (Li‐Cor's Biosciences).
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