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Invitrogen Ready Flow reagents are ready-to-use solutions designed to allow you to stain your cells for analysis by flow cytometry with:
Reach for the convenient dropper bottle, add 2 drops per 1 x 106 cells per milliliter of your single-cell suspension, and you're ready to incubate and analyze. Ready Flow reagents address your most common flow cytometry analysis needs, including dead cell identification and cell cycle (Figure 1) analysis. Preparing your samples for flow cytometry analysis is easy.
Figure 1. Cell cycle analysis with FxCycle Violet Ready Flow Reagent and Invitrogen Click-iT EdU Alexa Fluor 647 Flow Cytometry Assay Kit. Jurkat cells, a human T cell leukemia cell line, were pulsed with 10 µM EdU for 2 hours prior to detection with Alexa Fluor 647 azide. Cells were subsequently stained by adding 2 drops of FxCycle Violet Ready Flow Reagent and incubated for 30 minutes, at 25°C. Data was acquired on an Invitrogen Attune NxT Flow Cytometer using a 405 laser and 440/50 nm emission filter. Analysis of the population indicates the following distribution: apoptotic sub-G1 cells were 3.4%; G0/G1, 49.1%; S 33.0%; and G2M 14.0%.
SYTOX Green | Propidium Iodide | SYTOX AADvanced | TO-PRO 3 | DRAQ7 | |
---|---|---|---|---|---|
Readout | Dead cells become highly fluorescent following labeling with reagent, allowing for easy distinction from live cells; no calculations, no dilutions, no pipetting. | ||||
Mechanism of labeling | Enters cells upon loss of membrane integrity and binds to nucleic acids. | ||||
Target | Nucleic acids | Nucleic acids | Nucleic acids | dsDNA | dsDNA |
Fluorogenic | >100-fold fluorescence enhancement upon nucleic acid binding | Reagent is not fluorogenic; exhibits 20- to 30-fold fluorescence increase upon binding | >500-fold fluorescence enhancement upon nucleic acid binding | Reagent is not fluorogenic; exhibits fluorescence enhancement upon DNA binding | Reagent is not fluorogenic |
Storage | Room temperature | Room temperature | Room temperature | Room temperature | Room temperature |
Staining pattern fixable? | No | No | No | No | No |
Live-cell permeant? | No | No | No | No | No |
Ex/Em of reagent (in nm) | 504/523 | 535/617 | 546/647 | 642/661 | 488 to 647/677 |
Recommended excitation laser |
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Recommended bandpass filter |
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Cat. No. | R37168 | R37169 | R37173 | R37170 | D15107 |
Hoechst 33342 Ready Flow Reagent | FxCycle Violet Ready Flow Reagent | Vybrant DyeCycle Violet Ready Flow Reagent | |
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Readout | Fluorescent signal from reagent binding to DNA in live cells, allowing for cell cycle profiling and analysis; no calculations, no dilutions, no pipetting. | Fluorescent signal from reagent binding to DNA in fixed cells, allowing for cell cycle profiling and analysis; no calculations, no dilutions, no pipetting. | Fluorescent signal from reagent binding to DNA in live cells, allowing for cell cycle profiling and DNA content analysis; no calculations, no dilutions, no pipetting. |
Target | dsDNA | dsDNA | dsDNA |
Mechanism of labeling | The stain is a minor-groove DNA-binding reagent; binds preferentially to adenine–thymine regions of DNA. | Stain is cell permeable and, after binding double-stranded DNA, emits a fluorescent signal that is proportional to the DNA mass. | |
Fluorogenic | No. Fluorescence is greatly enhanced upon binding, but there is no modification of the dye structure. | ||
Storage | Room temperature | Room temperature | Room temperature |
Live-cell assay? | Yes | No | Yes |
Staining pattern fixable? | No | No | No |
Fixed-cell assay? | Yes | Yes | No |
Ex/Em of reagent (in nm) | 361/497 | 358/461 | 369/437 |
Recommended laser excitation | UV (350 nm) or 405 nm | UV (350 nm) or 405 nm | UV (350 nm) or 405 nm |
Recommended bandpass filter | 440/50 | 450/50 | 440/40 |
Cat. No. | R37165 | R37166 | R37172 |
Annexin V Pacific Blue Ready Flow Reagent | Annexin V Alexa Fluor 488 Ready Flow Reagent | Annexin V Alexa Fluor 647 Ready Flow Reagent | Annexin V APC Ready Flow Reagent | CellEvent Caspase-3/7 Green Ready Flow Kit | |
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Readout | Fluorescent signal from reagent binding to phosphatidyl serine in apoptotic cells; no calculations, no dilutions, no pipetting. | Fluorogenic substrate for detection of activated caspases 3 and 7 in apoptotic cells; no calculations, no dilutions, no pipetting. SYTOX AADvanced dead cell stain included in the kit for the detection of dead cells. | |||
Mechanism of labeling | In the presence of Ca2+, annexin V conjugates bind with high affinity to phosphatidylserine (PS), which becomes exposed on the outer leaflet of cells undergoing apoptosis. The annexin V conjugates can also pass through the compromised membranes of dead cells and bind to PS in the interior of the cell. Recommend using a cell-impermeant dead cell stain in combination with annexin V conjugate staining to distinguish dead cells from apoptotic cells. | The CellEvent reagent reagent consists of a four amino acid peptide (DEVD) conjugated to a nucleic acid binding dye. During apoptosis, caspase-3 and caspase-7 proteins are activated and are able to cleave the caspase 3/7 recognition sequence encoded in the DEVD peptide. | |||
Target | Binds to phosphatidylserine on apoptotic cell surfaces | Caspase 3/7 proteins, dead cells | |||
Fluorogenic | No | No | No | No | Yes; cleavage of the recognition sequence and binding of DNA by the reagent labels apoptotic cells with a bright, signal |
Storage | Room temperature | Room temperature | Room temperature | Room temperature | Room temperature |
Staining pattern fixable | No | No | No | No | No |
Live-cell permeant | No | No | No | No | Yes |
Ex/Em of reagent (in nm) | 410/455 | 495/519 | 650/665 | 650/660 | CellEvent Caspase-3/7: ~511/533; SYTOX AADvanced: 546/647 |
Recommended excitation laser | 405 nm | 488 nm | 633 or 635 nm | 633 or 635 nm | CellEvent Caspase-3/7: 488 nm; SYTOX AADvanced: 488 nm (blue), 532 nm (green) or 561 nm (yellow) |
Recommended bandpass filter | 450/50 | 530/30 | 661/8 | 661/8 | CellEvent Caspase-3/7: 530/30; SYTOX AADvanced: 695/40 |
Cat. No. | R37177 | R37174 | R37175 | R37176 | R37167 |
SYTOX Green | Propidium Iodide | SYTOX AADvanced | TO-PRO 3 | DRAQ7 | |
---|---|---|---|---|---|
Readout | Dead cells become highly fluorescent following labeling with reagent, allowing for easy distinction from live cells; no calculations, no dilutions, no pipetting. | ||||
Mechanism of labeling | Enters cells upon loss of membrane integrity and binds to nucleic acids. | ||||
Target | Nucleic acids | Nucleic acids | Nucleic acids | dsDNA | dsDNA |
Fluorogenic | >100-fold fluorescence enhancement upon nucleic acid binding | Reagent is not fluorogenic; exhibits 20- to 30-fold fluorescence increase upon binding | >500-fold fluorescence enhancement upon nucleic acid binding | Reagent is not fluorogenic; exhibits fluorescence enhancement upon DNA binding | Reagent is not fluorogenic |
Storage | Room temperature | Room temperature | Room temperature | Room temperature | Room temperature |
Staining pattern fixable? | No | No | No | No | No |
Live-cell permeant? | No | No | No | No | No |
Ex/Em of reagent (in nm) | 504/523 | 535/617 | 546/647 | 642/661 | 488 to 647/677 |
Recommended excitation laser |
|
|
|
|
|
Recommended bandpass filter |
|
|
|
|
|
Cat. No. | R37168 | R37169 | R37173 | R37170 | D15107 |
Hoechst 33342 Ready Flow Reagent | FxCycle Violet Ready Flow Reagent | Vybrant DyeCycle Violet Ready Flow Reagent | |
---|---|---|---|
Readout | Fluorescent signal from reagent binding to DNA in live cells, allowing for cell cycle profiling and analysis; no calculations, no dilutions, no pipetting. | Fluorescent signal from reagent binding to DNA in fixed cells, allowing for cell cycle profiling and analysis; no calculations, no dilutions, no pipetting. | Fluorescent signal from reagent binding to DNA in live cells, allowing for cell cycle profiling and DNA content analysis; no calculations, no dilutions, no pipetting. |
Target | dsDNA | dsDNA | dsDNA |
Mechanism of labeling | The stain is a minor-groove DNA-binding reagent; binds preferentially to adenine–thymine regions of DNA. | Stain is cell permeable and, after binding double-stranded DNA, emits a fluorescent signal that is proportional to the DNA mass. | |
Fluorogenic | No. Fluorescence is greatly enhanced upon binding, but there is no modification of the dye structure. | ||
Storage | Room temperature | Room temperature | Room temperature |
Live-cell assay? | Yes | No | Yes |
Staining pattern fixable? | No | No | No |
Fixed-cell assay? | Yes | Yes | No |
Ex/Em of reagent (in nm) | 361/497 | 358/461 | 369/437 |
Recommended laser excitation | UV (350 nm) or 405 nm | UV (350 nm) or 405 nm | UV (350 nm) or 405 nm |
Recommended bandpass filter | 440/50 | 450/50 | 440/40 |
Cat. No. | R37165 | R37166 | R37172 |
Annexin V Pacific Blue Ready Flow Reagent | Annexin V Alexa Fluor 488 Ready Flow Reagent | Annexin V Alexa Fluor 647 Ready Flow Reagent | Annexin V APC Ready Flow Reagent | CellEvent Caspase-3/7 Green Ready Flow Kit | |
---|---|---|---|---|---|
Readout | Fluorescent signal from reagent binding to phosphatidyl serine in apoptotic cells; no calculations, no dilutions, no pipetting. | Fluorogenic substrate for detection of activated caspases 3 and 7 in apoptotic cells; no calculations, no dilutions, no pipetting. SYTOX AADvanced dead cell stain included in the kit for the detection of dead cells. | |||
Mechanism of labeling | In the presence of Ca2+, annexin V conjugates bind with high affinity to phosphatidylserine (PS), which becomes exposed on the outer leaflet of cells undergoing apoptosis. The annexin V conjugates can also pass through the compromised membranes of dead cells and bind to PS in the interior of the cell. Recommend using a cell-impermeant dead cell stain in combination with annexin V conjugate staining to distinguish dead cells from apoptotic cells. | The CellEvent reagent reagent consists of a four amino acid peptide (DEVD) conjugated to a nucleic acid binding dye. During apoptosis, caspase-3 and caspase-7 proteins are activated and are able to cleave the caspase 3/7 recognition sequence encoded in the DEVD peptide. | |||
Target | Binds to phosphatidylserine on apoptotic cell surfaces | Caspase 3/7 proteins, dead cells | |||
Fluorogenic | No | No | No | No | Yes; cleavage of the recognition sequence and binding of DNA by the reagent labels apoptotic cells with a bright, signal |
Storage | Room temperature | Room temperature | Room temperature | Room temperature | Room temperature |
Staining pattern fixable | No | No | No | No | No |
Live-cell permeant | No | No | No | No | Yes |
Ex/Em of reagent (in nm) | 410/455 | 495/519 | 650/665 | 650/660 | CellEvent Caspase-3/7: ~511/533; SYTOX AADvanced: 546/647 |
Recommended excitation laser | 405 nm | 488 nm | 633 or 635 nm | 633 or 635 nm | CellEvent Caspase-3/7: 488 nm; SYTOX AADvanced: 488 nm (blue), 532 nm (green) or 561 nm (yellow) |
Recommended bandpass filter | 450/50 | 530/30 | 661/8 | 661/8 | CellEvent Caspase-3/7: 530/30; SYTOX AADvanced: 695/40 |
Cat. No. | R37177 | R37174 | R37175 | R37176 | R37167 |
Cy™ is a trademark or registered trademark of GE Healthcare.
Not for resale. Super Bright Polymer Dyes are sold under license from Becton, Dickinson and Company.
Brilliant Ultra Violet™ and Brilliant Violet™ are trademarks or registered trademarks of Becton, Dickinson and Company or its affiliates, and are used under license.
For Research Use Only. Not for use in diagnostic procedures.