BD Horizon Brilliant™ Violet 480 Imaging Reagents · 2014-09-22 · BD Biosciences has...
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BD Biosciences has exclusively developed BD Horizon Brilliant™ Violet 480 (BV480) reagents to enable imaging applications. Part of the family of unique polymer dyes acquired from Sirigen Ltd., BV480 dye is brighter than conventional dyes and can be used in combination with most common imaging dyes, including Alexa Fluor® 488, Alexa Fluor® 555, Alexa Fluor® 594, Alexa Fluor® 647, DRAQ5™ and DRAQ7™, DAPI, and BD Horizon™ BV421. Using BV480 along with Alexa Fluor® 488, Alexa Fluor® 555, Alexa Fluor® 647, and DAPI, expands the number of parameters that can be analyzed in an imaging application. This can provide better signal resolution due to significantly brighter and spectrally distinct dyes when used in combination with the appropriate filter configuration (Figure 1, Figure 2, and Table 1). Moreover, as a bright dye, BV480 provides a high level of signal, enabling shorter exposure times than standard imaging fluorophores (such as Alexa Fluor® 488) (Figure 3). BV480 is also compatible with standard fixation and permeabilization buffers, as well as ProLong® Gold antifade embedding medium, which results in excellent photostability (Figure 4). The BD Horizon Brilliant™ unique polymer dyes were developed from pioneering polymer dye technology acquired from Sirigen Ltd. These dyes deliver more and brighter dye choices over conventional dyes with equivalent background—a breakthrough in the field that enables increased sensitivity. Features Very bright dye that expands color choice for multicolor imaging Minimal spillover into DAPI and GFP, or Alexa Fluor® 488 Greater sensitivity in comparison to traditional fluorescent dyes Photostable For Research Use Only. Not for use in diagnostic or therapeutic procedures. Visit bdbiosciences.com for more information. Figure 1. Multicolor immunocytochemistry using BV480 Cells: MCF-7 cells. Fix: BD Cytofix™ Fixation Buffer (Cat. No. 554655). Wash/ Perm: 0.1% Triton™ X-100. Block/Stain: 5% goat serum, 1% BSA, 0.5% Triton X-100 in 1X PBS. Antibodies: Purified Mouse Anti-E-Cadherin (Cat. No. 610182) with BD Horizon™ BV480 Goat anti-Mouse (Cat. No. 564877); Alexa Fluor® 488 Mouse anti-β-Tubulin (Cat No. 558605), Alexa Fluor® 555 Mouse anti-Human Ki-67 (Cat. No. 558617), Alexa Fluor® 647 Mouse anti-GM130 (Cat. No. 558712). Other reagents: DAPI (Cat. No. 564907), ProLong® Gold antifade (Life Technologies). Instrument: BD Pathway™ 435 Cell Analyzer and merged using BD Attovision™ Software (BD Biosciences). 20x objective. Table 1. Filters used in this study BD Horizon Brilliant ™ Violet 480 Imaging Reagents Dye Excitation (nm) Dichroic Mirror (nm) Emission (nm) BV480 445/20 475 485/20 Alexa Fluor® 488 490/30 506 537/26 Alexa Fluor® 555 543/22 562 593/40 DRAQ5 628/40 660 692/40 BV421 390/18 409 434/17 DAPI Tubulin Alexa Fluor® 488 GM130 Alexa Fluor® 647 E Cadherin BV480 Ki67 Alexa Fluor® 555 Merge For additional instrument configurations, visit bdbiosciences.com/research/cellularimaging.
Transcript of BD Horizon Brilliant™ Violet 480 Imaging Reagents · 2014-09-22 · BD Biosciences has...
BD Biosciences has exclusively developed BD Horizon Brilliant™ Violet 480 (BV480) reagents to enable imaging applications. Part of the family of unique polymer dyes acquired from Sirigen Ltd., BV480 dye is brighter than conventional dyes and can be used in combination with most common imaging dyes, including Alexa Fluor® 488, Alexa Fluor® 555, Alexa Fluor® 594, Alexa Fluor® 647, DRAQ5™ and DRAQ7™, DAPI, and BD Horizon™ BV421.
Using BV480 along with Alexa Fluor® 488, Alexa Fluor® 555, Alexa Fluor® 647, and DAPI, expands the number of parameters that can be analyzed in an imaging application. This can provide better signal resolution due to significantly brighter and spectrally distinct dyes when used in combination with the appropriate filter configuration (Figure 1, Figure 2, and Table 1).
Moreover, as a bright dye, BV480 provides a high level of signal, enabling shorter exposure times than standard imaging fluorophores (such as Alexa Fluor® 488) (Figure 3).
BV480 is also compatible with standard fixation and permeabilization buffers, as well as ProLong® Gold antifade embedding medium, which results in excellent photostability (Figure 4).
The BD Horizon Brilliant™ unique polymer dyes were developed from pioneering polymer dye technology acquired from Sirigen Ltd. These dyes deliver more and brighter dye choices over conventional dyes with equivalent background—a breakthrough in the field that enables increased sensitivity.
Features
Very bright dye that expands color choice for multicolor imaging
Minimal spillover into DAPI and GFP, or Alexa Fluor® 488
Greater sensitivity in comparison to traditional fluorescent dyes
Photostable
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Visit bdbiosciences.com for more information.
Figure 1. Multicolor immunocytochemistry using BV480
Cells: MCF-7 cells. Fix: BD Cytofix™ Fixation Buffer (Cat. No. 554655). Wash/Perm: 0.1% Triton™ X-100. Block/Stain: 5% goat serum, 1% BSA, 0.5% Triton X-100 in 1X PBS. Antibodies: Purified Mouse Anti-E-Cadherin (Cat. No. 610182) with BD Horizon™ BV480 Goat anti-Mouse (Cat. No. 564877); Alexa Fluor® 488 Mouse anti-β-Tubulin (Cat No. 558605), Alexa Fluor® 555 Mouse anti-Human Ki-67 (Cat. No. 558617), Alexa Fluor® 647 Mouse anti-GM130 (Cat. No. 558712). Other reagents: DAPI (Cat. No. 564907), ProLong® Gold antifade (Life Technologies). Instrument: BD Pathway™ 435 Cell Analyzer and merged using BD Attovision™ Software (BD Biosciences). 20x objective.
Table 1. Filters used in this study
BD Horizon Brilliant™ Violet 480 Imaging Reagents
Dye Excitation (nm) Dichroic Mirror (nm) Emission (nm)
BV480 445/20 475 485/20
Alexa Fluor® 488 490/30 506 537/26
Alexa Fluor® 555 543/22 562 593/40
DRAQ5 628/40 660 692/40
BV421 390/18 409 434/17
DAPI
Tubulin Alexa Fluor® 488
GM130 Alexa Fluor® 647
E Cadherin BV480
Ki67 Alexa Fluor® 555
Merge For additional instrument configurations, visit bdbiosciences.com/research/cellularimaging.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.
Alexa Fluor® and ProLong® are registered trademarks of Life Technologies Corporation.
DRAQ5 and DRAQ7 are trademarks of BioStatus Limited.
Triton is a trademark of Sigma-Aldrich.
BD, BD Logo and all other trademarks are property of Becton, Dickinson and Company. © 2014 BD
23-16728-00
BD Horizon Brilliant™ Violet 480 Imaging Reagents
Description Clone Size Cat.No.
Goat anti-mouse Ig Poly 1270 50 µg 564877
Goat anti-Rat Ig Poly 1271 50 µg 564878
Streptavidin 0.1 mg 564876
BD Horizon BV480 Reagents
BD Biosciencesbdbiosciences.com
Figure 3. Immunohistochemistry using BV480
Cells: Frozen human colon. Fix: BD Cytofix Fixation Buffer (Cat. No. 554655). Perm: 0.25% Triton X-100. Block/Stain: 5% goat serum, 1% BSA, 0.2% gelatin, 0.2% Triton X in PBS. Wash: 0.2% Triton X-100. Antibodies: Purified Mouse Anti-E-Cadherin (Cat. No. 610182) with BD Horizon BV480 Goat anti-Mouse (Cat No. 564877) or Alexa Fluor® 488 Goat Anti-Mouse (Life Technologies). Other reagents: DRAQ5 (BioStatus), ProLong® Gold antifade (Life Technologies). Instrument: BD Pathway 435 Cell Analyzer and merged using BD Attovision Software (BD Biosciences). 20x objective.
Figure 4. Photostability of BV480: continuous exposure to excitation light
Cells: U2OS cells. Fix, perm, and blocking: BD Cytofix Fixation Buffer (Cat. No. 554655). Antibodies: Purified Mouse anti-Human CD324 (Cat No. 562869) with BD Horizon BV480 Goat anti-Mouse (Cat. No.564877) or Alexa Fluor® 488 Goat anti-Mouse (Life Technologies). Other reagents: ProLong® Gold antifade (Life Technologies). Instrument: BD Pathway 435 Cell Analyzer (BD Biosciences). 20x objective.
Figure 2. Compatibility of BV480 with BD Horizon™ BV421 for immunofluorescent imaging
Cells: H9 human embryonic stem cells (WiCell, Madison WI). Fix: BD Cytofix Fixation Buffer (Cat. No. 554655). Perm: BD Phosflow™ Perm Buffer III (Cat. No. 558050). Block/Stain: 5% goat serum, 1% BSA, 0.5% Triton X-100 in 1X PBS. Wash: 0.1% Triton X-100. Antibodies: Purified Mouse anti-Oct3/4a (Cat. No. 561555) with BD Horizon BV480 Goat Anti-Mouse (Cat. No.564877), BD Horizon BV421 Mouse Anti-Human TRA-1-60 (Cat. No. 562711). Other reagents: DRAQ5 (BioStatus), ProLong® Gold antifade (Life Technologies). Instrument: BD Pathway 435 Cell Analyzer and merged using BD Attovision Software (BD Biosciences). 20x objective.
Oct 3/4a BV480
A
DRAQ5
BV4800.07 s
Time: 0 2 m 4.5 m
Alexa Fluor® 4880.13 s
C
Tra-1-60 BV421
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Merge
D
BV480 0.015 s
A
Alexa Fluor® 488 0.14 s
B
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Alexa Fluor® 488 BV480
