Brilliant Violet dye conjugated secondary antibodies from Jackson ImmunoResearch allow the addition of more colors to multiple labeling assays. With appropriate filters, Brilliant Violet 421 (BV421) and Brilliant Violet 480 (BV480) provide two options, with emission in the violet and blue channels respectively. Effective five-color fluorescent labeling is possible when BV421 and BV480 are combined with Alexa Fluor 488, Rhodamine Red-X, and Alexa Fluor 647 conjugates. If nuclear counterstaining is desired, four-color antibody staining is possible using BV421, BV480, Alexa Fluor 488, and Rhodamine Red-X. Switching the nuclear stain from DAPI (4’,6-diamidino-2-phenylindole; emission in the blue region) to DRAQ5 (deep red anthraquinone 5, which has red emission) frees the violet–blue region of the spectrum to accommodate the two Brilliant Violet dyes. DRAQ5’s excitation and emission profiles overlap those of Alexa Fluor 647. BV421 and BV480 are available conjugated to a range of secondary antibodies recommended for multiple labeling due to their minimal cross-reactivity.
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