Rabbit anti drosophila myc

Total proteins were isolated from Drosophila tissues and the cultured cells with different treatments, and the protein concentration in the lysates was quantified using Bio-Rad protein assay reagent. Equal amounts of total protein were subjected to western blotting. The antibodies and dilutions used in the study were as follows: goat anti-Drosophila Fzr (1:1000; Santa Cruz), rabbit anti-Drosophila Myc (1:1000; Santa Cruz), goat anti-human Fzr (1:1000; Santa Cruz), mouse anti-human Myc (1:1000; Santa Cruz), mouse anti-Drosophila CycB (1:5000; DSHB), rabbit anti-human CycB (1:5000; Cell Signaling), mouse anti-H2B (1:10 000; Beyotime), rabbit anti-H2Bub (1:20 000; Cell Signaling), mouse anti-V5 (1:5000; Abcam), rabbit anti-Flag (1:5000; Sigma), mouse anti-Myc tag (1:5000; Sigma), rabbit anti-MCM6 (1:5000; Zoonbio Biotechnology), and mouse anti-tubulin (1:10 000; Beyotime). The following secondary antibodies were used, including HRP-conjugated goat anti-rabbit (1:10 000; Beyotime), goat anti-mouse (1:10 000; Beyotime), and donkey anti-goat (1:10 000; Beyotime).

Immunostaining assay was performed in the salivary glands and cultured cells as previously described (35 (link),45 (link)). Briefly, fixed cells or tissues were washed three times with PBST buffer (1× PBS including 0.3% Triton-X 100) and then stained at 4°C overnight with primary antibodies at the following dilutions: goat anti-Drosophila Fzr (1:50; Santa Cruz), rabbit anti-Drosophila Myc (1:50; Santa Cruz), goat anti-human Fzr (1:50; Santa Cruz), mouse anti-human Myc (1:50; Santa Cruz), mouse anti-CycB (1:200; DSHB), rabbit anti-MCM6 (1:200; Zoonbio Biotechnology) and mouse anti-Myc tag (1:200; Sigma). After washing three times with PBST buffer, the samples were incubated with the corresponding Alexa Fluor-conjugated secondary antibodies (Life Technologies). DAPI (1:1000; Thermo Fisher Scientific) was used for nuclear labeling, while Alexa Fluor-conjugated phalloidin (1:1000; Life Technologies) was used for Actin staining. Finally, after washing with PBS three times, the cells or tissues were mounted in Vectashield mounting buffer and the fluorescence signals were captured by confocal microscope (Zeiss LSM 880 and Olympus Fv1000).