Comparison of enhanced chemiluminescence (ECL) and WesternDot® fluorescence–based western blot protein detection. Serial dilutions (750–3 ng protein) of GAPDH were run on NuPAGE® Novex® 4–12% Bis-Tris precast gels (Cat. No. NP0321BOX) and transferred to iBlot® nitrocellulose membranes (Cat. No. IB3010-32) using the iBlot® Gel Transfer Device (Cat. No. IB1001). The membranes were probed with anti-GAPDH antibodies followed by either ECL detection using a horseradish peroxidase goat anti–mouse IgG or fluorescence detection using WesternDot® 655 goat anti–mouse IgG (Cat. No. W10813). Images were collected using the Fujifilm® LAS- 4000 gel imager. Partial blot images show GAPDH detection, and the graph shows the detection sensitivities of the two techniques.
|Tested species reactivity||Mouse|
|Host / Isotype||Goat / IgG|
|Storage buffer||50mM borate, pH 8.3, with 1M betaine|
|Contains||0.05% sodium azide|
|Storage Conditions||4° C, do not freeze|
|Antibody Form||Whole Antibody|
|Tested Applications||Dilution *|
|Western Blot (WB)||1:500|
* 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.
WesternDot fluorescently labeled secondary antibodies deliver superior signal-to-noise ratios and exhibit comparable sensitivity to enhanced chemiluminescence (ECL)-based methods in western blot applications. No stripping and re-probing or multiple blots are required for multiple protein detection as commonly needed with ECL or colorimetric detection. Differently labeled WesternDot antibodies can be applied to a single blot for simultaneous detection of multiple proteins using standard gel or blot imaging platforms (a list of instruments that are compatible with WesternDot detection can be found in the user manual below). Our WesternDot antibodies utilize Qdot nanocrystals enhanced with VIVID technology, making them brighter than the original Qdot reagents. Additionally, they have reduced intensity differences between the different colors, simplifying multiplex applications.
Anti-Mouse secondary antibodies are affinity-purified antibodies with well-characterized specificity for mouse 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.