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 Goat anti-Rat IgG (H+L) Cross-Adsorbed Secondary Antibody, DSB-X Biotin (Product # D-20697) at dilutions 1:2,000 (Fig. 1), 1:10,000 (Fig. 2) and 1:20,000 (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|
|Published species reactivity||Not Applicable|
|Host / Isotype||Goat / IgG|
|Immunogen||Gamma Immunoglobins Heavy and Light chains|
|Storage buffer||PBS, pH 7.5|
|Contains||5mM sodium azide|
|Storage Conditions||4° C, store in dark|
|Cross Adsorption||Against human serum and mouse IgG and serum|
|Antibody Form||Whole Antibody|
|Tested Applications||Dilution *|
|Immunofluorescence (IF)||1-10 µg/mL|
|Western Blot (WB)||1:2,000-1:20,000|
* 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.
DSB-X biotin is a derivative of desthiobiotin, a stable biotin percursuor. DSB-X biotin utilizes a seven-atom spacer to increase the available of the DSB-X biotin conjugate to bind in the deep biotin-binding pocket of streptpavidin or avidin. Whereas harsh chaotropic agents and low pH are required to dissociate a biotin complex from avidin or steptavidin, binding can be reversed by adding excess D-desthiobiotin or natural D-biotin at neutral pH and at room temperature.
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.
Keratinocyte-derived chemokine induces prostate epithelial hyperplasia and reactive stroma in a novel transgenic mouse model.
D-20697 was used in immunohistochemistry to test if overexpressed IL-8 and keratinocyte-derived chemokine induce prostate epithelial hyperplasia and a reactive phenotype
|Schauer IG,Ressler SJ,Rowley DR||The Prostate (69:373)||2009|
|Not Applicable||Not Cited||Single-molecule analysis of cadherin-mediated cell-cell adhesion.||Panorchan P,Thompson MS,Davis KJ,Tseng Y,Konstantopoulos K,Wirtz D||Journal of cell science (119:66)||2006|
|Not Applicable||Not Cited||Easily reversible desthiobiotin binding to streptavidin, avidin, and other biotin-binding proteins: uses for protein labeling, detection, and isolation.||Hirsch JD,Eslamizar L,Filanoski BJ,Malekzadeh N,Haugland RP,Beechem JM,Haugland RP||Analytical biochemistry (308:343)||2002|