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ReadiUse™ Preactivated PE Maleimide [Activated R-Phycoerythrin]

Flow cytometry analysis of whole blood cells stained with CD8 (SK1)-PE conjugate prepared with ReadiUse™ Preactivated PE Maleimide (Cat No. 2565). The fluorescence signal was monitored using an Aurora spectral flow cytometer in the B4-A channel.
Flow cytometry analysis of whole blood cells stained with CD8 (SK1)-PE conjugate prepared with ReadiUse™ Preactivated PE Maleimide (Cat No. 2565). The fluorescence signal was monitored using an Aurora spectral flow cytometer in the B4-A channel.
Flow cytometry analysis of whole blood cells stained with CD8 (SK1)-PE conjugate prepared with ReadiUse™ Preactivated PE Maleimide (Cat No. 2565). The fluorescence signal was monitored using an Aurora spectral flow cytometer in the B4-A channel.
Flow cytometry analysis of whole blood stained with PE anti-human CD4 *SK3* conjugate. The fluorescence signal was monitored using an Aurora spectral flow cytometer in the PE specific B4-A channel.
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Physical properties
Spectral properties
Correction Factor (280 nm)0.175
Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)565
Emission (nm)574
Quantum yield0.82
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageRefrigerated (2-8 °C); Minimize light exposure; Desiccated
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See also: PE and APC
Correction Factor (280 nm)
Extinction coefficient (cm -1 M -1)
Excitation (nm)
Emission (nm)
Quantum yield
ReadiUse™ Preactivated PE Maleimide is prepared by reacting highly purified R-Phycoerythrin (PE) with SMCC. The NHS group of SMCC reacts with the lysine groups of PE, leaving maleimide groups available to react with free sulfhydryl groups on proteins to be conjugated. ReadiUse™ Preactivated PE Maleimide is highly purified to completely remove the unreacted SMCC, and lyophilized to a powder form. It is ready to use and will conjugate without further preparation upon mixing with sulfhydryl-containing target molecules such as reduced IgGs. This activated phycobiliprotein can be easily conjugated to antibodies and other proteins without the use of added chemical crosslinking agents. The prepared conjugates maintain the spectral characteristics of PE.

Example protocol


Reduction of IgG
  1. Prepare a fresh solution of 1.0 M DTT (15.4 mg/100 µL) in distilled water. Antibody reduction can be carried out in different buffers, such as MES, phosphate, or TRIS (pH range 6 to 8). For best results, antibody solutions should be >2 mg/mL. If less than 2 mg/mL, the antibody should be concentrated.
  2. 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 temperature for 30 minutes without additional mixing to minimize the reoxidation of cysteines.
  3. Purify the reduced antibody using a desalting column (Cat No. 60500) pre-equilibrated with 50 mM MES Buffer (pH 6.0-6.5) and 2 mM EDTA.
  4. Measure the antibody concentration using a Nanodrop. (Con. (mg/mL) = A280nm/1.4). Note: Reduced antibodies are typically unstable. It is best to run the conjugation reaction immediately following purification. 

Conjugate with ReadiUse™ Preactivated PE Maleimide
  1. Reconstitute ReadiUse™ Preactivated PE Maleimide in ddH2O to make a 10 mg/mL solution. For Cat No. 2565 (1 mg) reconstitute in 100 µL of ddH2O. If using Cat No. 2566 (5 mg), reconstitute in 500 µL of ddH2O. Reconstituted ReadiUse™ Preactivated PE Maleimide solutions are stable at 4 °C for up to one week, protected from light.
  2. Add the reduced antibody directly to the solution of ReadiUse™ Preactivated PE at a ratio of 250-300 µg PE/100 µg of reduced antibody.
  3. Rotate the mixture for 60 to 120 minutes at room temperature.
  4. Block any free sulfhydryl groups on the antibody. Prepare a fresh solution of 10 mg/mL N-Ethylmaleimide (NEM) in DMSO and add 3.4 µL per mg of antibody. Rotate for 20 minutes at room temperature. 

Conjugate Purification
  1. Purify the antibody/PE conjugate solution using size exclusion chromatography. Note: Typically the antibody/PE conjugate can be stored at 2-8 °C, protected from light, for up to 6 months. The best storage conditions for any particular conjugate must be determined by experimentation and depends on the stability of the antibody. 


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Spectral properties

Correction Factor (280 nm)0.175
Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)565
Emission (nm)574
Quantum yield0.82

Product Family

NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)Correction Factor (280 nm)
ReadiUse™ Preactivated APC Maleimide [Activated Allophycocyanin]6516607300000.195
ReadiUse™ Preactivated PerCP Maleimide4776784060000.22



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Difference between mitogen-stimulated B and T cells in nonspecific binding of R-phycoerythrin-conjugated antibodies.
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Journal: Journal of immunoassay & immunochemistry (2018): 84-98
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Authors: Kim, Myun Soo and Kim, Tae Sung
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2013): 452-60
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Authors: Trajanoski, Daniel and Fidler, Samantha J
Journal: Methods in molecular biology (Clifton, N.J.) (2012): 47-65
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