Rabbit monoclonal antibody to digoxigenin

ELISA to detect the amount of PMO in muscle tissues was performed as described elsewhere.⁴⁸ (link) In brief, DNA probe was designed with sequences complementary to PMO (synthesized by The Beijing Genomics Institute, Beijing, China) as follows: 3′-CCGGTTTGGAGCCGAATGGACTTTA-5′, with phosphorothioate ends in bold. The 5′ and 3′ ends of the probes were labeled with digoxigenin and biotin, respectively. Standard PMO samples and muscle tissues (100 mg/mL) were digested with 20 mg/mL proteinase K at 55°C overnight. Following PMO-probe hybridization, the avidin-biotin interaction of the hybridized probe was performed on Pierce NeutrAvidin Coated 96-well plates, Black (Thermo Fisher Scientific, MA, USA). Unhybridized probes were digested with micrococcal nuclease at 10 U/μL (Thermo Fisher Scientific, MA, USA). Then the hybridized probes were reacted with rabbit monoclonal antibody to digoxigenin (Cell Signaling Technology, MA, USA), followed by detection with peroxidase-conjugated goat anti-rabbit IgG (Abcam, Cambridge, UK). Signals from the PMO-hybridized probe were detected at 450 nm with TME Substrate (Solarbio, Beijing, China) in a monochromator EnSpire Multimode plate reader (PerkinElmer, Boston, MA, USA).