Simultaneous detection of three proteins on a single blot using Qdot® secondary antibody conjugates. A western blot containing serial dilutions (20–3 µg protein) of lysates from unstimulated (lanes 2–5) and hEGF-stimulated (lanes 6–9) A431 cells was probed with mouse anti-EGFR, rabbit anti–phospho-EGFR, and chicken anti-GAPDH antibodies, followed by (A) WesternDot® 800 goat anti-mouse (pseudocolored blue), (B) WesternDot® 605 goat anti-rabbit (pseudocolored red), and (C) WesternDot® 655 goat anti-chicken (pseudocolored green) conjugates. (D) The merged image shows overlaid red and blue bands as purple. The blot contains MagicMark™ XP Western Protein Standard (lane 1, Cat. No. LC5603) and was imaged using the Fujifilm® LAS-4000 gel imager.
|Tested species reactivity||Mouse|
|Host / Isotype||Donkey / 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.