Figure 2. IHC analysis of NPHS2 using anti- NPHS2 antibody (PA1322-1). NPHS2 was detected in frozen section of rat kidney tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1ug/ml rabbit anti- NPHS2 Antibody (PA1322-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

Figure 3. Western blot analysis of NPHS2 using anti- NPHS2 antibody (PA1322-1). 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 50ug of sample under reducing conditions. Lane 1: human 293T whole cell lysates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 70 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti- NPHS2 antigen affinity purified polyclonal antibody (Catalog # PA1322-1) at 0.5 ug/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 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for NPHS2 at approximately 45KD. The expected band size for NPHS2 is at 42KD.