Western blot analysis was performed on whole cell extracts (30 µg lysate) of A549 (Lane 1) and MKN45 (Lane 2). The blots were probed with Anti-Alpha Tubulin Rat Monoclonal Antibody (Product # MA1-80017, 1 µg/ml) and detected by chemiluminescence using Rabbit anti-Rat IgG (H+L) Secondary Antibody, Biotin (Product # 31834) at dilutions 1:50,000 (Fig. 1), 1:2,00,000 (Fig. 2) and 1:4,00,000 (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||Rabbit / IgG|
|Storage buffer||PBS, pH 7.8, with 3mg/ml BSA|
|Contains||0.08% sodium azide|
|Storage Conditions||4° C|
|Tested Applications||Dilution *|
|ELISA (ELISA)||1:20,000 - 1:400,000|
|Flow Cytometry (Flow)||1:200 - 1:1,000|
|Immunocytochemistry (ICC)||1:500 - 1:5,000|
|Immunofluorescence (IF)||1:500 - 1:5,000|
|Immunohistochemistry (IHC)||1:500 - 1:5,000|
|Immunoprecipitation (IP)||1:500 - 1:5,000|
|Western Blot (WB)||1:50,000-1:400,000|
* 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.
Product # 31834 has been successfully used in Western blot, IF, ICC, IHC, IP and FACS applications.
Product # 31834 reacts with heavy chains of rat IgG and with the light chains common to most rat immunoglobulins. Based on solid-phase enzyme immunoassay, this antibody cross-reacts with horse IgG, hamster IgG, and mouse IgG (12%), bovine IgG, mouse IgM, swine IgG, and guinea pig IgG (6%), cat IgG, goat IgG, Rhesus monkey IgG, sheep IgG, and swine IgG (3%). It has minimum cross-reactivity with immunoglobulins from other species.
Store product at 4°C until opened. To extend the shelf-life of this product, add an equal volume of glycerol to make a final concentration of approximately 50% glycerol and store at -20°C.
Reconstitute with 1.0 ml of distilled water (1.5 mg/ml after restoration).
Thermo Scientific 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.