Western Blot Analysis of Viral and Host Proteins
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Corresponding Organization :
Other organizations : The University of Texas Medical Branch at Galveston, New York University
Variable analysis
- Cell lysates were resolved on 7.5% Mini-Protean TGX SDS-PAGE gels
- Membranes were blocked with 5% (wt/vol) nonfat dry milk in TBST (TBS plus 0.1% [vol/vol] Tween 20) for 1 h
- Membranes were probed with the indicated primary antibody in 3% (wt/vol) bovine serum albumin in TBST at 4°C overnight
- Membranes were probed with the following secondary antibodies in 5% (wt/vol) nonfat dry milk in TBST for 1 h at room temperature
- Viral and host protein analysis
- Cell lysates were transferred to polyvinylidene difluoride membranes using a Trans-Blot Turbo transfer system (Bio-Rad)
- Proteins were visualized using ECL or SuperSignal West Femto chemiluminescence reagents (Pierce) and detected by autoradiography
- Anti-pSTAT1 Y701 (1:1,000; Cell Signaling Technologies, 9171L)
- Anti-STAT1 D1K9Y (1:1,000; Cell Signaling Technologies, 14994P)
- Anti-IFITM1 (1:1,000; Invitrogen, PA5-20989)
- Anti-SARS-CoV/CoV-2 Spike 1A9 (1:1,000; GeneTex, GTX632604)
- Anti-β-actin (1:1,000; Abcam, ab8227)
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