Fluorescent Detection, Immobilon-FL Transfer Membrane, Bløk-FL Noise Cancelling Reagents

Fluorescent Detection

Fluorescence-based detection of Western blots, while increasing in popularity due to multiplex detection capabilities, requires specialized tools to obtain optimal results. The reagents presented here have been optimized to work together for fast, reproducible fluorescent Westerns.

Successful fluorescence detection of Western blots depends on the following components:

Transfer membrane - High protein binding capacity with low autofluorescence gives the best signal to noise ratio.
Blocking reagents - Protein free blockers eliminate the non-specific background mediated by protein blockers.
Antibody specificity - Purified antibodies are more likely to produce less non-specific background.
Fluorophore used - Some fluorophores are sticky by nature, while others photobleach easily. Protect the blot at all times with light-proof material, such as aluminum foil.

Immobilon-FL Transfer Membrane

Click to Enlarge.

This 0.45 μm membrane is the first transfer membrane specifically optimized for fluorescencebased detection of Western blots. Its extremely low background autofluorescence improves sensitivity of all fluorescence detection protocols.

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Key Benefits of Immobilon-FL Transfer Membrane:

The only membrane that works at near-infrared wavelengths (700-800nm)
Strong signals due to higher protein adsorption & retention on the membrane
Low background to detect even faint bands
High tensile strength for multiple stripping and reprobing cycles

Bløk^™-FL Noise Cancelling Reagents

Fluorescent Blok noise cancelling reagents

Bløk-FL reagent provides the best signal-to-noise results. Dilution series of EGF-stimulated A431 lysate (2-0.5 µg/lane,12-110) were resolved by SDS-PAGE and transferred onto Immobilon-FL membranes after equilibration of the gel for 30 minutes. Click to Enlarge.

This chemical-based, protein-free blocker decreases background caused by non-specific antibody binding without leaving a thick, sticky layer similar to milk.

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Key Benefits of Blok-FL Noise Cancelling Reagents:

Specially formulated for reduced background on fluorescent Westerns
Ready to use straight from the bottle
Stable at room temperature for 2 years
Enables colorimetric staining of the blots after immunodetection