Super Bright Polymer Dyes
Antibody conjugates for the violet laser
Invitrogen eBioscience Super Bright dyes are a series of fluorophores that are based on a fluorescent polymer and its tandems, named for their emission wavelength and excited by the violet laser (405 nm). These dyes are optimized for use in flow cytometry and may allow for better discrimination of dim populations due to their brightness. Super Bright antibody conjugates increase your options for multicolor panel design and allow you to expand the utility of your violet laser.
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Super Bright 436 dye
Invitrogen eBioscience Super Bright 436 dye has an excitation maximum of 414 nm and an emission peak of 436 nm. A 450/50 bandpass filter or equivalent is recommended, similar to Invitrogen™ eFluor™ 450 dye or Invitrogen Pacific Blue dye. Super Bright 436 dye–conjugated antibodies are significantly brighter than those conjugated to eFluor 450 dye, and are an alternative to Brilliant Violet 421 conjugates with similar resolution of positive and negative populations. Stability studies indicate that Super Bright 436 dye exhibits a minimal loss of fluorescence when stained cells are exposed to formaldehyde fixative for up to three days or left uncovered overnight in ambient light.
Figure 1. Super Bright 436 dye performance comparison. Mouse bone marrow cells were stained with Anti-Ly-6A/E (clone D7) APC and either Anti-CD117 (clone 2B8) conjugated to (left panel) Super Bright 436 dye, (middle panel) eFluor 450 dye or (right panel) Brilliant Violet 421 dye.
Super Bright 600 dye
Invitrogen eBioscience Super Bright 600 dye is a tandem dye comprising Super Bright 436 dye and an acceptor dye that emits at 600 nm. It can be detected using a 610/20 bandpass filter. Antibodies conjugated to this tandem polymer dye are comparable in brightness to Brilliant Violet 605 dye conjugates. Super Bright 600 dye–conjugated antibodies are stable for up to three days when stored in a formaldehyde fixative solution.
Figure 2. Multiplex flow cytometry experiment. Normal human peripheral blood cells were diluted in Super Bright Staining Buffer, then surface stained with the indicated reagents. Viable cells (determined using a fixable viability stain) were used for analysis to discriminate various T cell subsets.
Super Bright Staining Buffer
As with other polymer dyes, nonspecific dye–dye interactions may form when multiple Super Bright dye–conjugated antibodies are used together in the same experiment. When they occur, these interactions can cause unexpected population shifts. Use of the Super Bright Staining Buffer (Cat. No. SB-4400-42) prior to addition of Super Bright conjugates alleviates this issue and returns populations to their expected position within the plot. When using Super Bright dyes in combination with other polymer dyes, such as Brilliant Violet dyes, the Super Bright Staining Buffer can be used to minimize dye–dye interaction. Super Bright Staining Buffer is formulated to be used at 5 µL/test, making it convenient for use when preparing cocktails.
Figure 3. Super Bright Staining Buffer mitigates non-specific polymer dye interactions. (A) Flow Cytometry Staining Buffer (top plot) or Super Bright Staining Buffer (bottom plot) was added to human peripheral blood cells prior to staining with Anti-CD8a (clone RPA-T8) Super Bright 600 dye and Anti-CD4 (clone SK3) Super Bright 436 dye. (B) Flow Cytometry Staining Buffer (top plot) or Super Bright Staining Buffer (bottom plot) was added to human peripheral blood cells prior to staining with Anti-CD8a (clone SK1) Brilliant Violet 605 dye and Anti-CD4 (clone SK3) Super Bright 436 dye. (C) Flow Cytometry Staining Buffer (top plot) or Super Bright Staining Buffer (bottom plot) was added to human peripheral blood cells prior to staining with Anti-CD8a (clone RPA-T8) Super Bright 600 dye and Anti-CD4 (clone SK3) Brilliant Violet 421 dye.
|Dye||Super Bright 436||Super Bright 600|
|Excitation laser (nm)||Violet
|Suggested bandpass filter||450/50||610/20|
|Application||Flow cytometry||Flow cytometry|
|Comparable to||Brilliant Violet 421||Brilliant Violet 605|
|Primary antibodies||Super Bright 436 conjugates||Super Bright 600 conjugates|
|Control antibodies||Super Bright 436 conjugates||Super Bright 600 conjugates|
|Super Bright Staining Buffer (Cat. No. SB-4400-42) is recommended for use with Super Bright polymer dyes.|
For Research Use Only. Not for use in diagnostic procedures.