Rat anti rna pol 2

Samples were incubated overnight at 4°C with the following primary antibodies: mouse anti‐m3G (1:50; Santa Cruz Biotechnology, https://www.scbt.com), mouse anti‐U2B″ (1:20; LS Bio, https://www.lsbio.com) and rat anti‐RNA pol II (elongated form, hyperphosphorylated Ser2 of the C‐terminal domain; 1:100; Chromotek, https://www.chromotek.com) in 0.01% acBSA in PBS, pH 7.2. After rinsing with PBS, the samples were incubated for 1 h at 37°C with the following secondary antibodies: anti‐mouse Alexa 546 (ThermoFisher Scientific), anti‐rat Alexa 488 (ThermoFisher Scientific) or anti‐rabbit Alexa 488 (ThermoFisher Scientific) 1:500 in 0.01% acBSA in PBS, pH 7.2. Next, the slides were rinsed with PBS, pre‐incubated with 2% BSA in PBS, pH 7.2, for 15 min, and incubated overnight at 4°C with the anti‐Sm ANA No. 5 antibody diluted 1:300 in 0.2% acBSA in PBS, pH 7.2. After rinsing with PBS, the samples were incubated for 1 h at 37°C with the appropriate respective anti‐human antibody: Alexa 488 at 1:750 or TRITC, at 1:200 in 0.01% acBSA in PBS, pH 7.2.