Vio® Bright V423 fluorophore

Brightest Vio Dye for the violet laser
Allows confident detection of rare cells as well as dim and uncharacterized markers
Minimal compensation effort
Exclusive collection of REAfinity™ Recombinant Antibody conjugates

Vio Bright V423 is Miltenyi Biotec’s brightest dye excited at 405 nm. In addition to high brightness this dye shows a significantly improved signal-to-noise ratio, minimal intra-laser compensation and a superior detection of rare antigens.
Vio Bright V423 is a synthetic dye which is 5-times brighter compared to the VioBlue® Dye. This makes Vio Bright V423 an excellent choice for multiparameter cell analysis.

Fluorophore at a glance

Overview of fluorophore characteristics

Laser	405 nm
Excitation max	399 nm
Emission max	420 nm
Application	Flow cytometry
Common filter set (FC)	450/50 nm
Stain index	5 out of 5
Photostablility	Photostable to ambient light
Fixation stability	PFA

Laser and filter compatibility

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Absorption and emission spectra of Vio Bright V423

Vio Bright V423 can be excited with a violet laser (405 nm) and displays peak excitation and emission at 399 nm and 423 nm, respectively. Its fluorescence signal can be detected with a standard 450/50 nm filter set from all major flow cytometry hardware providers, for example, MACSQuant® Flow Cytometers. Both absorption and emission spectra of Vio Bright V423 are very similar to the spectra of polymer dyes Brilliant Violet 421 and Super Bright 436.

Enhanced brightness

Compared to polymer-based fluorochromes with high fluorescence intensities, such as Brilliant Violet 421 and Super Bright 436, Vio Bright V423 exhibits a similar or brighter degree of fluorescence. Increased brightness of Vio Bright V423 conjugates is observed across different antigens; the data below show examples of CD19 and CD56 staining.

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Higher specificity of fluorescence signal

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Superior detection of rare antigens due to high brightness of Vio Bright V423

Vio Bright V423 shows exceptional brightness allowing confident detection of rare cells as well as dim and uncharacterized markers. Vio Bright V423 provides excellent resolution for the analysis of rare antigens such as CD25, which makes it a perfect fit for multiparameter analysis. Note that Vio Bright V423 gives even brighter signals than Vio Bright B515, which is one of our brightest Vio Dyes.

Decreased spillover

Vio Bright V423 requires only minimal compensation

Compared to the other dyes excited by violet lasers, Vio Bright V423 shows minimal spillover into the neighboring detection channels. Thus, with a minimal change in compensation settings, Vio Bright V423 provides the benefit of a brighter dye.

Compensation matrix for a CD19 antibody conjugated to different fluorochromes excited by a violet laser. Values depict the percentage of compensation required for the respective detection filter.

	Conjugate	V1-A (450/50nm)	V2-A (525/50 nm)	V3-A (579/34 nm)	V4-A (615/20 nm)	V5-A (667/30 nm)
V1-A (450/50 nm)	CD19-Vio Bright V423	100	4	1
V2-A (525/50 nm)	CD19-BV510	40	100	60	40	6
V3-A (579/34 nm)	CD19-BV570	40	3	100	65	10
V5-A (667/30 nm)	CD19-BV650	30		3	52	100

Fixation stability

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Excellent performance with different staining protocols

Vio Bright V423 shows excellent stability after fixation with paraformaldehyde and is compatible with different surface and intracellular staining protocols and buffer sets. Conjugate performance may vary depending on antigen nature upon fixation.

Photostability

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Comparable photostability over time

The stability of Vio Bright V423 upon exposure to ambient light was analyzed at different time points. Analysis of the photostability of CD19-Vio Bright V423 indicated a minor drop in both MFI and SI after up to 24 hours of continuous exposure to ambient light (~850 lux), which is similar compared to commercially available alternatives.

Resources for Vio Bright V423

Multicolor flow cytometry panels

Miltenyi Biotec–tested panels (MBTPs) are a collection of multicolor panels that are optimized and pre-designed for specific flow cytometry applications. They are developed and validated by Miltenyi Biotec’s R&D experts, as well as by the customers who actually use Miltenyi Biotec antibodies.

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