ReadiUse™ Preactivated PE-Cy7 Maleimide
Ordering information
| Price | |
| Catalog Number | |
| Unit Size | |
| Quantity |
Additional ordering information
| Telephone | 1-800-990-8053 |
| Fax | 1-800-609-2943 |
| sales@aatbio.com | |
| International | See distributors |
| Bulk request | Inquire |
| Custom size | Inquire |
| Shipping | Standard overnight for United States, inquire for international |
Physical properties
| Molecular weight | N/A |
| Solvent | Water |
Spectral properties
| Extinction coefficient (cm -1 M -1) | 1960000 |
| Excitation (nm) | 565 |
| Emission (nm) | 778 |
Storage, safety and handling
| H-phrase | H303, H313, H333 |
| Hazard symbol | XN |
| Intended use | Research Use Only (RUO) |
| R-phrase | R20, R21, R22 |
| Storage | Refrigerated (2-8 °C); Minimize light exposure |
Related products
| Overview |  SDSProtocol |
See also: PE and APC
Molecular weight N/A | Extinction coefficient (cm -1 M -1) 1960000 | Excitation (nm) 565 | Emission (nm) 778 |
PE-Cy7 is a popular tandem color used in flow cytometry. Its primary absorption peak is at 565 nm with emission peak at ~780 nm. AAT Bioquest offers this preactivated PE-Cy7 maleimide to facilitate the PE-Cy7 tandem conjugations to antibodies and other proteins such as streptavidin and other secondary reagents. It is prepared from the commonly used crosslinker SMCC. Our preactivated PE-Cy7 maleimide is ready to conjugate the reduced antibodies and other thiol-containing biomolecules.
Example protocol
SAMPLE EXPERIMENTAL PROTOCOL
Reduction of IgG
- Prepare a fresh solution of 1.0 M DTT (15.4 mg/100 µL) in distilled water. Antibody solutions should be at 2 mg/mL or higher for best results. The reduction can be carried out in different buffers for example: MES, phosphate, and TRIS buffers (pH range 6 to 8). The antibody should be concentrated if less than 2 mg/mL.
- Add 2 µL of 1.0 M DTT stock per 100 µL of antibody solution and mix well. Let the antibody solution stand at room temp for 30 minutes without additional mixing (to minimize reoxidation of cysteines to cystines).
- Purify the reduced antibody over a desalting column pre-equilibrated with 50 mM MES Buffer (pH=6.0-6.5) with 2 mM EDTA. (Desalting column: https://www.aatbio.com/products/readiuse-bio-gel-p-6-spin-column?unit=60500)
- Measure the Antibody concentration with Nanodrop. (Con. (mg/ml)= A280nm/1.4) Note: The reduced antibody is not stable; the conjugation reaction needs to be carried out the conjugation as soon as possible after purification.
Conjugate with ReadiUse™ Preactivated PE-Cy7 Maleimide
- Reconstitute ReadiUse™ Preactivated PE-Cy7 Maleimide in 100 µL ddH2O to 10 mg/mL. Note: Reconstituted ReadiUse™ Preactivated PE-Cy7 Maleimide are stable at 4 °C for one week, please kept it from light.
- Add reduced antibody to pre-activated PE-Cy7 directly to at the ratio of 250-300 µg PE-Cy7 /100 µg reduced antibody.
- Rotate the mixture for 60-120 mins at room temperature.
- After 60 minutes, block the free sulfhydryls on the antibody.
- Prepare a fresh solution of 10 mg/mL NEM in DMSO; add 3.4 µL per mg of antibody and rotate for 20 minutes at room temperature.
Purification
- Antibody/PE-Cy7 conjugate could be further purified through size exclusion chromatography to get best performance. Note: The Antibody/PE-Cy7 conjugate solution is recommended to be stored at 2~8 °C and kept from light.
Spectrum
Open in Advanced Spectrum Viewer
Spectral properties
| Extinction coefficient (cm -1 M -1) | 1960000 |
| Excitation (nm) | 565 |
| Emission (nm) | 778 |
Product Family
| Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) |
| ReadiUse™ Preactivated PE-Cy7 Tandem | 565 | 778 | 1960000 |
| ReadiUse™ Preactivated PE-Cy5 Maleimide | 565 | 666 | 1960000 |
| ReadiUse™ Preactivated APC-Cy7 Maleimide | 651 | 779 | 700000 |
| ReadiUse™ Preactivated PerCP-Cy7 Maleimide | 482 | 784 | 350000 |
Images
Figure 1. Flow cytometry analysis of whole blood stained with PE-Cy7 anti-human CD4 *SK3* conjugate. The fluorescence signal was monitored using an Aurora flow cytometer in the PE-Cy7 specific B13-A channel.
References
View all 37 references: Citation Explorer
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