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Tech Tips and Protocols

Practical advice and lab tips from Biotium scientists on using fluorescence in your research

Tech tip: Imaging bacteria using agarose pads

Bacteria can be more difficult to image than other cell types because they are small, usually non-adherent, and are often motile. Here we describe how to image live or fixed bacteria using agarose pads to immobilize the cells during the imaging process, allowing you to perform multi-color fluorescence microscopy experiments.

Tech Tip: Battling Tissue Autofluorescence

Autofluorescence in tissues from lipofuscin, red blood cells, and extracellular matrix can make it all but impossible to distinguish specific fluorescence staining from non-specific background. See our protocols for quenching autofluorescence with TrueBlack®.

Tech tip: Avoiding Artifacts from UV Photoconversion of DAPI and Hoechst

In multi-color imaging experiments, limiting cross-talk between detection channels is crucial for obtaining specific results. Many researchers are unaware that photoconversion of DAPI and Hoechst can cause nuclear stains to bleed into the FITC channel. Learn about the different ways to avoid it.

Tech Tip: Multi-color fluorescence imaging using Biotium’s Tyramide Amplification Kits

Tyramide signal amplification (TSA) is an immunofluorescence technique that uses enzymatic coupling to label targets with large amounts of dye, resulting in dramatically higher signal. Here, we provide protocols for performing multiple TSA reactions on the same sample.

Tech Tip: Combined Direct and Indirect Immunofluorescence Using Primary Antibodies from the Same Host

Multicolor staining using fluorescent secondary antibodies provides signal amplification for greater sensitivity, but requires that primary antibodies be raised in different hosts. Directly labeled primaries from the same host can used simultaneously, but signal can suffer. A combination of indirect and direct methods takes advantage of the best of both for added flexibility in multicolor staining. See our protocol to learn more.

Protocols for antibody-based detection

Commonly used protocols for performing immunofluorescence staining for microscopy, flow cytometry, and western blotting.