mFluor™ Green 630 SE
Ordering information
Price | |
Catalog Number | |
Unit Size | |
Quantity |
Additional ordering information
Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
Quotation | Request |
International | See distributors |
Shipping | Standard overnight for United States, inquire for international |
Physical properties
Molecular weight | 806.94 |
Solvent | DMSO |
Spectral properties
Extinction coefficient (cm -1 M -1) | 510001 |
Excitation (nm) | 537 |
Emission (nm) | 657 |
Storage, safety and handling
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
Storage | Freeze (< -15 °C); Minimize light exposure |
UNSPSC | 12171501 |
Alternative formats
mFluor™ Blue 630 SE |
mFluor™ Yellow 630 SE |
Overview | ![]() ![]() |
See also: Amine Reactive Dyes and Probes for Conjugation, Bioconjugation, Chemical Reagents, Dyes by Functional Group, Flow Cytometry Reagents, mFluor™ Dyes and Kits, Spectral Flow Cytometry
Molecular weight 806.94 | Extinction coefficient (cm -1 M -1) 510001 | Excitation (nm) 537 | Emission (nm) 657 |
Advances in spectral flow cytometers have expanded applications and capabilities beyond conventional flow cytometry. Now with spectral flow cytometry analysis, researchers and scientists can investigate an increasing number of molecules of interest. However, the potential of spectral flow cytometry is severely limited by the availability of fluorescent labels and readouts. AAT Bioquest's mFluor™ dyes are developed for multicolor flow cytometry-focused applications, in particular, for spectral fluorescence flow cytometry. mFluor™ Green 630 dye can be well excited with green laser at 532 nm. It has a huge Stokes shift with emission ~630 nm. mFluor™ Green 630 dyes are water-soluble, and the protein conjugates prepared with mFluor™ Green 630 dyes are well excited at 532 nm to give red fluorescence. mFluor™ Green 630 dye and conjugates are excellent green laser reagents for flow cytometry detections. Compared to RPE, mFluor™ Green 630 dyes are much more photostable, making them readily available for fluorescence imaging applications while it is very difficult to use RPE conjugates for fluorescence imaging applications due to the rapid photobleaching of RPE conjugates. It is also a unique fluorochrome for spectral flow cytometry since there are very few existing dyes that have this spectral profile.
Example protocol
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.
Note The pH of the protein solution (Solution A) should be 8.5 ± 0.5. If the pH of the protein solution is lower than 8.0, adjust the pH to the range of 8.0-9.0 using 1 M sodium bicarbonate solution or 1 M pH 9.0 phosphate buffer.
Note The protein should be dissolved in 1X phosphate buffered saline (PBS), pH 7.2-7.4. If the protein is dissolved in Tris or glycine buffer, it must be dialyzed against 1X PBS, pH 7.2-7.4, to remove free amines or ammonium salts (such as ammonium sulfate and ammonium acetate) that are widely used for protein precipitation.
Note Impure antibodies or antibodies stabilized with bovine serum albumin (BSA) or gelatin will not be labeled well. The presence of sodium azide or thimerosal might also interfere with the conjugation reaction. Sodium azide or thimerosal can be removed by dialysis or spin column for optimal labeling results.
Note The conjugation efficiency is significantly reduced if the protein concentration is less than 2 mg/mL. For optimal labeling efficiency the final protein concentration range of 2-10 mg/mL is recommended.
Note Prepare the dye stock solution (Solution B) before starting the conjugation. Use promptly. Extended storage of the dye stock solution may reduce the dye activity. Solution B can be stored in freezer for two weeks when kept from light and moisture. Avoid freeze-thaw cycles.
1. Protein stock solution (Solution A)
Mix 100 µL of a reaction buffer (e.g., 1 M sodium carbonate solution or 1 M phosphate buffer with pH ~9.0) with 900 µL of the target protein solution (e.g. antibody, protein concentration >2 mg/mL if possible) to give 1 mL protein labeling stock solution.Note The pH of the protein solution (Solution A) should be 8.5 ± 0.5. If the pH of the protein solution is lower than 8.0, adjust the pH to the range of 8.0-9.0 using 1 M sodium bicarbonate solution or 1 M pH 9.0 phosphate buffer.
Note The protein should be dissolved in 1X phosphate buffered saline (PBS), pH 7.2-7.4. If the protein is dissolved in Tris or glycine buffer, it must be dialyzed against 1X PBS, pH 7.2-7.4, to remove free amines or ammonium salts (such as ammonium sulfate and ammonium acetate) that are widely used for protein precipitation.
Note Impure antibodies or antibodies stabilized with bovine serum albumin (BSA) or gelatin will not be labeled well. The presence of sodium azide or thimerosal might also interfere with the conjugation reaction. Sodium azide or thimerosal can be removed by dialysis or spin column for optimal labeling results.
Note The conjugation efficiency is significantly reduced if the protein concentration is less than 2 mg/mL. For optimal labeling efficiency the final protein concentration range of 2-10 mg/mL is recommended.
2. mFluor™ Green 630 SE stock solution (Solution B)
Add anhydrous DMSO into the vial of mFluor™ Green 630 SE to make a 10 mM stock solution. Mix well by pipetting or vortex.Note Prepare the dye stock solution (Solution B) before starting the conjugation. Use promptly. Extended storage of the dye stock solution may reduce the dye activity. Solution B can be stored in freezer for two weeks when kept from light and moisture. Avoid freeze-thaw cycles.
SAMPLE EXPERIMENTAL PROTOCOL
This labeling protocol was developed for the conjugate of Goat anti-mouse IgG with mFluor™ Green 630 SE. You might need further optimization for your particular proteins.
Note Each protein requires distinct dye/protein ratio, which also depends on the properties of dyes. Over labeling of a protein could detrimentally affects its binding affinity while the protein conjugates of low dye/protein ratio gives reduced sensitivity.
Note Each protein requires distinct dye/protein ratio, which also depends on the properties of dyes. Over labeling of a protein could detrimentally affects its binding affinity while the protein conjugates of low dye/protein ratio gives reduced sensitivity.
Run conjugation reaction
- Use 10:1 molar ratio of Solution B (dye)/Solution A (protein) as the starting point: Add 5 µL of the dye stock solution (Solution B, assuming the dye stock solution is 10 mM) into the vial of the protein solution (95 µL of Solution A) with effective shaking. The concentration of the protein is ~0.05 mM assuming the protein concentration is 10 mg/mL and the molecular weight of the protein is ~200KD.
Note We recommend to use 10:1 molar ratio of Solution B (dye)/Solution A (protein). If it is too less or too high, determine the optimal dye/protein ratio at 5:1, 15:1 and 20:1 respectively. - Continue to rotate or shake the reaction mixture at room temperature for 30-60 minutes.
Purify the conjugation
The following protocol is an example of dye-protein conjugate purification by using a Sephadex G-25 column.- Prepare Sephadex G-25 column according to the manufacture instruction.
- Load the reaction mixture (From "Run conjugation reaction") to the top of the Sephadex G-25 column.
- Add PBS (pH 7.2-7.4) as soon as the sample runs just below the top resin surface.
- Add more PBS (pH 7.2-7.4) to the desired sample to complete the column purification. Combine the fractions that contain the desired dye-protein conjugate.
Note For immediate use, the dye-protein conjugate need be diluted with staining buffer, and aliquoted for multiple uses.
Note For longer term storage, dye-protein conjugate solution need be concentrated or freeze dried.
Calculators
Common stock solution preparation
Table 1. Volume of DMSO needed to reconstitute specific mass of mFluor™ Green 630 SE to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 123.925 µL | 619.625 µL | 1.239 mL | 6.196 mL | 12.392 mL |
5 mM | 24.785 µL | 123.925 µL | 247.85 µL | 1.239 mL | 2.478 mL |
10 mM | 12.392 µL | 61.962 µL | 123.925 µL | 619.625 µL | 1.239 mL |
Molarity calculator
Enter any two values (mass, volume, concentration) to calculate the third.
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Spectrum
Open in Advanced Spectrum Viewer


Spectral properties
Extinction coefficient (cm -1 M -1) | 510001 |
Excitation (nm) | 537 |
Emission (nm) | 657 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Correction Factor (260 nm) | Correction Factor (280 nm) |
mFluor™ Blue 630 SE | 470 | 634 | 490001 | 0.197 | 0.275 |
mFluor™ Yellow 630 SE | 570 | 632 | 1100001 | - | - |
Images

Figure 1. Fluorescent dye NHS esters (or succinimidyl esters) are the most popular tool for conjugating dyes to a peptide, protein, antibody, amino-modified oligonucleotide, or nucleic acid. NHS esters react readily with the primary amines (R-NH2) of proteins, amine-modified oligonucleotides, and other amine-containing molecules. The resulting dye conjugates are quite stable.
References
View all 19 references: Citation Explorer
A Comprehensive Workflow for Applying Single-Cell Clustering and Pseudotime Analysis to Flow Cytometry Data.
Authors: Melsen, Janine E and van Ostaijen-Ten Dam, Monique M and Lankester, Arjan C and Schilham, Marco W and van den Akker, Erik B
Journal: Journal of immunology (Baltimore, Md. : 1950) (2020)
Authors: Melsen, Janine E and van Ostaijen-Ten Dam, Monique M and Lankester, Arjan C and Schilham, Marco W and van den Akker, Erik B
Journal: Journal of immunology (Baltimore, Md. : 1950) (2020)
High-Dimensional Data Analysis Algorithms Yield Comparable Results for Mass Cytometry and Spectral Flow Cytometry Data.
Authors: Ferrer-Font, Laura and Mayer, Johannes U and Old, Samuel and Hermans, Ian F and Irish, Jonathan and Price, Kylie M
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
Authors: Ferrer-Font, Laura and Mayer, Johannes U and Old, Samuel and Hermans, Ian F and Irish, Jonathan and Price, Kylie M
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
HTLV infected individuals have increased B-cell activation and proinflammatory regulatory T-cells.
Authors: Kjerulff, Bertram and Petersen, Mikkel Steen and Rodrigues, Candida Medina and da Silva Té, David and Christiansen, Mette and Erikstrup, Christian and Hønge, Bo Langhoff
Journal: Immunobiology (2020): 151878
Authors: Kjerulff, Bertram and Petersen, Mikkel Steen and Rodrigues, Candida Medina and da Silva Té, David and Christiansen, Mette and Erikstrup, Christian and Hønge, Bo Langhoff
Journal: Immunobiology (2020): 151878
Multispectral Flow Cytometry: Unaddressed Issues and Recommendations for Improvement.
Authors: Parks, David R
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
Authors: Parks, David R
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
High-dimensional analysis of intestinal immune cells during helminth infection.
Authors: Ferrer-Font, Laura and Mehta, Palak and Harmos, Phoebe and Schmidt, Alfonso J and Chappell, Sally and Price, Kylie M and Hermans, Ian F and Ronchese, Franca and le Gros, Graham and Mayer, Johannes U
Journal: eLife (2020)
Authors: Ferrer-Font, Laura and Mehta, Palak and Harmos, Phoebe and Schmidt, Alfonso J and Chappell, Sally and Price, Kylie M and Hermans, Ian F and Ronchese, Franca and le Gros, Graham and Mayer, Johannes U
Journal: eLife (2020)
Panel Design and Optimization for High-Dimensional Immunophenotyping Assays Using Spectral Flow Cytometry.
Authors: Ferrer-Font, Laura and Pellefigues, Christophe and Mayer, Johannes U and Small, Sam J and Jaimes, Maria C and Price, Kylie M
Journal: Current protocols in cytometry (2020): e70
Authors: Ferrer-Font, Laura and Pellefigues, Christophe and Mayer, Johannes U and Small, Sam J and Jaimes, Maria C and Price, Kylie M
Journal: Current protocols in cytometry (2020): e70
Phenotypic Analysis of the Mouse Hematopoietic Hierarchy Using Spectral Cytometry: From Stem Cell Subsets to Early Progenitor Compartments.
Authors: Solomon, Michael and DeLay, Monica and Reynaud, Damien
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
Authors: Solomon, Michael and DeLay, Monica and Reynaud, Damien
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)
Acquisition of High-Quality Spectral Flow Cytometry Data.
Authors: Fox, Amy and Dutt, Taru S and Karger, Burton and Obregón-Henao, Andrés and Anderson, G Brooke and Henao-Tamayo, Marcela
Journal: Current protocols in cytometry (2020): e74
Authors: Fox, Amy and Dutt, Taru S and Karger, Burton and Obregón-Henao, Andrés and Anderson, G Brooke and Henao-Tamayo, Marcela
Journal: Current protocols in cytometry (2020): e74
Spectral flow cytometry-Quo vadimus?
Authors: Robinson, J Paul
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2019): 823-824
Authors: Robinson, J Paul
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2019): 823-824
Time-Delayed Integration-Spectral Flow Cytometer (TDI-SFC) for Low-Abundance-Cell Immunophenotyping.
Authors: Hu, Wenting and Soper, Steven A and Jackson, J Matt
Journal: Analytical chemistry (2019): 4656-4664
Authors: Hu, Wenting and Soper, Steven A and Jackson, J Matt
Journal: Analytical chemistry (2019): 4656-4664
Application notes
A New Protein Crosslinking Method for Labeling and Modifying Antibodies
Abbreviation of Common Chemical Compounds Related to Peptides
Bright Tide Fluor™-Based Fluorescent Peptides and Their Applications In Drug Discovery and Disease Diagnosis
FITC (Fluorescein isothiocyanate)
Fluorescein isothiocyanate (FITC)
Abbreviation of Common Chemical Compounds Related to Peptides
Bright Tide Fluor™-Based Fluorescent Peptides and Their Applications In Drug Discovery and Disease Diagnosis
FITC (Fluorescein isothiocyanate)
Fluorescein isothiocyanate (FITC)