Goat anti rabbit 680cw

Stage 21HH and 24HH legs were homogenised in RIPA buffer (Fisher) containing protease inhibitors, centrifuged, and the supernatant is collected. Protein concentration was estimated using a DC Protein Assay kit (Bio-Rad). Recombinant mouse SHH N-terminus protein (R&D Systems) was used as a positive control. Protein samples were loaded as individual limbs per lane. Proteins were separated by electrophoresis using pre-cast 12% gels (Invitrogen), transferred to nitrocellulose membranes by standard procedures. Membranes were blocked in Odyssey Blocking Buffer (Licor), incubated with 1:100 rabbit anti-SHH H-160 (Santa Cruz, sc9024) 1:2500 mouse anti-γ-tubulin (Sigma, T5326) 4 °C overnight, followed by goat anti-rabbit 680CW (Licor, 926-32221) goat anti-mouse 800CW (Licor, 926-32210) for 1 hour. Membranes were dried and signal detected using an Odyssey Infrared Imager (Licor). Bands were quantified using Image Studio software, and normalised to γ-tubulin protein.

Primary mouse antibodies to proteins IE4 (HR-VZV-20), ORF9p (HR-VZV-38), ORF23p (HR-VZV-12), and IE63 (HR-VZV-33) were acquired commercially from Center for Proteomics, University of Rijeka (CapRi). gE (SC-56995) was acquired from Santa Cruz Biotechnology (Dallas, TX, USA) and β-actin (A00702) was purchased from GeneScript (Piscataway, NJ, USA). Rabbit antibody to α-Tubulin (600-401-880) was purchased from Rockland Immunochemicals (Limerick, PA, USA). Rabbit antibodies to IE62, ORF29p, and IE4 were reported previously [53 (link)–55 (link)].
Secondary IRDye antibodies for immunoblotting compatible with the LI-COR Odyssey were purchased from LI-COR: goat anti-mouse 680CW (926–32220), goat anti-mouse 800CW (926–32210), goat anti-rabbit 680CW (926–32221), goat anti-rabbit 800CW (926–32211). Those used for immunohistochemistry were purchased from Invitrogen: Alexa Fluor 488 goat anti-mouse (A11029), Alexa Fluor 546 goat anti-mouse (A11030), Alexa Fluor 488 goat anti-rabbit (A11034), Alexa Fluor 546 goat anti-rabbit (A11035).