Western blot analysis was performed on whole cell extracts (30 µg lysate) of A549 (Lane 1) and F9 (Lane 2). The blots were probed with Anti-alpha Tubulin Rat Monoclonal Antibody (Product# MA1-80017, 1 µg/ml) and detected by chemiluminescence using Donkey anti-Rat IgG (H+L) Secondary Antibody, Biotin (Product # A18743) at dilutions 1:10,000 (Fig. 1), 1:20,000 (Fig. 2) and 1:50,000 (Fig. 3). A 52 kDa band corresponding to alpha Tubulin was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary antibody after blocking with 5 % skimmed milk. This is followed by incubating the membrane with Poly-HRP Streptavidin (Product# N200, 1:10,000 dilution). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
|Tested species reactivity||Rat|
|Host / Isotype||Donkey / IgG|
|Immunogen||Gamma Immunoglobins Heavy and Light chains|
|Storage buffer||PBS, pH 7.2, with 1% BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.|
|Antibody Form||Whole Antibody|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1:500-1:20,000|
|Western Blot (WB)||1:10,000-1:50,000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
The sensitivity of each lot of antibody is confirmed using ELISA. The specificity of each lot of antibody is confirmed by isoelectric focusing (IEF).
Anti-Rat secondary antibodies are affinity-purified antibodies with well-characterized specificity for rat immunoglobulins and are useful in the detection, sorting or purification of its specified target. Secondary antibodies offer increased versatility enabling users to use many detection systems (e.g. HRP, AP, fluorescence). They can also provide greater sensitivity through signal amplification as multiple secondary antibodies can bind to a single primary antibody. Most commonly, secondary antibodies are generated by immunizing the host animal with a pooled population of immunoglobulins from the target species and can be further purified and modified (i.e. immunoaffinity chromatography, antibody fragmentation, label conjugation, etc.) to generate highly specific reagents.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.