Nb100 59787

Primary antibodies specific for HBV proteins included anti-HBV core protein mouse monoclonal antibody (MAb) 8C9-11 (72 (link)), anti-HBV core protein mouse MAb sc-23945 (Santa Cruz), both recognizing linear epitopes within the HBV core protein, and HBV capsid/capsid-intermediate-specific rabbit polyclonal antibody (PAb) B0586 (Dako). Primary antibodies used for epitope tags and cellular targets included anti-HA rat MAb 3F10 (Core Facility for Monoclonal Antibodies, Helmholtz Zentrum München), mouse MAb against the His₆ tag (Clontech), anti-PML protein mouse MAb sc-966 (Santa Cruz), polyclonal rabbit anti-PML protein PAb NB100-59787 (Novus Biologicals), polyclonal rabbit anti-PML protein ab72137 (Abcam), monoclonal mouse anti-SUMO2/3 ab81371 (Abcam), and anti-beta-actin mouse MAb AC-15 (Sigma-Aldrich). Secondary antibodies conjugated to Alexa Fluor 488 were purchased from Thermo Scientific, and secondary antibodies coupled to horseradish peroxidase or Alexa Fluor 647 were anti-rabbit IgG, anti-mouse IgG, anti-mouse light chain IgG, and anti-rat IgG (all from Dianova).

Whole-cell protein lysates were prepared using RIPA lysis buffer as recently described (46 (link)). Lysates were separated by SDS-PAGE, transferred on a nitrocellulose blotting membrane (0.45 µm), and detected via western blotting as previously described (88 (link)). As primary antibody for HBV Core protein, mouse mAb 8C9-11 (71 (link)), mouse mAb sc-23945 (Santa Cruz), and HBV Capsid rabbit pAb B0586 (DAKO) were used. Cellular protein steady-state levels were determined using polyclonal rabbit Ab raised against PML protein (NB100-59787; Novus Biologicals), and β-actin mouse mAb AC-15 as a control (Sigma-Aldrich, Inc.). mAb3F10 was used for the detection of HA-tagged proteins. To detect proteins by immunoblotting, secondary anti-rabbit IgG and anti-mouse IgG antibodies conjugated to horseradish peroxidase (Jackson/Dianova) were used.
Western blots were processed using Adobe Photoshop CS5 and Adobe Illustrator CS5 software. For relative quantification of protein steady-state levels and comparisons, ImageJ 1.52a (89 (link)) was used.