Immunohistochemistry analysis of alpha Actinin 3 on paraffin-embedded mouse skeletal muscle tissue. Antigen retrieval was performed using citrate buffer (pH6, epitope retrieval solution) for 20 mins. Sample was blocked using 10% goat serum, incubated with alpha Actinin 3 polyclonal antibody (Product# PA5-78720) with a dilution of 1 µg/mL (overnight at 4°C), and followed by biotinylated goat anti-rabbit IgG (30 minutes at 37°C). Development was performed using Streptavidin-Biotin-Complex (SABC) with DAB chromogen method.

Western blot analysis of ACTN3/Alpha Actinin 3 in, Lane 1: human HT1080 whole cell lysates, Lane 2: rat skeletal muscle tissue lysates, Lane 3: rat skeletal muscle tissue lysates, Lane 4: mouse skeletal muscle tissue lysates, Lane 5: mouse skeletal muscle tissue lysates. Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 µg of sample under reducing conditions. After Electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. The membrane was blocked with 5% non-fat milk/TBS for 1. 5 hour at RT. The membrane was incubated with alpha Actinin 3 Polyclonal Antibody (Product # PA5-78720) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0. 1% Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5,000 for 1. 5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit. A specific band was detected for ACTN3/Alpha Actinin 3 at approximately 103 kDa. The expected band size for ACTN3/Alpha Actinin 3 is at 103 kDa.