Application Notes: Western blot, 0.1-0.5µg/ml Immunohistochemistry (Paraffin-embedded Section), 0.5-1µg/ml ELISA (Cap), 1-5µg/ml. Add 0.2ml of distilled water will yield a concentration of 500ug/ml
IHC analysis of VCAM1 using anti-VCAM1 antibody. VCAM1 was detected in paraffin-embedded section of mouse spleen tissue. 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 1 ug/ml rabbit anti-VCAM1 Antibody overnight at 4 Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.
IHC analysis of VCAM1 using anti-VCAM1 antibody. VCAM1 was detected in paraffin-embedded section of rat spleen tissue. 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 1 ug/ml rabbit anti-VCAM1 Antibody overnight at 4 Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.
Sandwich ELISA - Recombinant mouse VCAM1 protein standard curve. Use in combination with reagents from Mouse VCAM1 ELISA Kit EZ-Set (DIY Antibody Pairs).
Western blot analysis of VCAM1 using anti-VCAM1 antibody. 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 50 ug of sample under reducing conditions. Lane 1: mouse spleen tissue lysates, Lane 2: mouse lung tissue lysates, Lane 3: rat spleen tissue lysates, Lane 4: rat lung tissue lysates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-VCAM1 antigen affinity purified polyclonal antibody at 0.5 ug/mL overnight at 4 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 with Tanon 5200 system. A specific band was detected for VCAM1 at approximately 110KD. The expected band size for VCAM1 is at 81KD.
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