ReadiUse™ Preactivated PE

R-Phycoerythrin (PE) is isolated from red algae. Its primary absorption peak is at 565 nm with secondary peaks at 496 and 545 nm. AAT Bioquest offers this preactivated PE to facilitate the PE conjugations to antibodies and other proteins such as streptavidin and other secondary reagents. Our preactivated PE is ready to conjugate, giving much higher yield than the conventionally tedious SMCC-based conjugation chemistry. In addition, our preactivated PE is conjugated to a protein via its amino group that is abundant in proteins while SMCC chemistry targets the thiol group that has to be regenerated by the reduction of antibodies.

Example protocol

AT A GLANCE

Important PE was premodified with our Buccutite™ FOL. Your antibody (or other proteins) is modified with our Buccutite™ MTA to give MTA-modified protein. The MTA-modified protein readily reacts with FOL-modified PE (provided) to give the desired PE-antibody conjugate.

SAMPLE EXPERIMENTAL PROTOCOL

Preparation of pre-activated Antibody with Buccutite™ MTA

Reconstitute Buccutite™ MTA in DMSO at ~10 mg/mL.
Note Store unused MTA at -20 °C; it can be used for up to two freeze and thaw cycles.
Prepare target antibody (Ab) in pH = 8.5 - 9.0 buffer at a concentration above 1 mg/ml.
Add the MTA to Ab solution at the ratio of 8 - 10 µg MTA/100 µg Ab.
Mix well and react at room temperature for 60 minutes, rotating during the reaction.
Purify the reaction mixture with a desalting column to remove any unreacted MTA. Exchange the buffer to PBS or another buffer of your choice.
Collect the MTA-activated Ab. Estimate the concentration by 70% yield of the original starting amount.

Conjugate with Pre-activated PE

Reconstitute pre-activated PE in 100 µL ddH₂O to 10 mg/mL.
Note Reconstituted pre-activated PE is not stable and can not be stored for more than one month.
Add pre-activated PE directly to MTA-activated target Ab solution at the ratio of 300 µg PE/100 µg MTA-activated Ab.
Rotate the mixture for 1 - 2 hours at room temperature.
The Ab/PE conjugates are now ready to use.
Note The antibody conjugate should be stored at >0.5 mg/mL in the presence of a carrier protein (e.g., 0.1% bovine serum albumin) and 0.02-0.05% sodium azide.
Note The Ab/PE can be stored at 4 °C for two months.
Optional: Ab/PE can be further purified through size exclusion chromatography to get better performance.

Spectrum

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Product family

Name	Excitation (nm)	Emission (nm)	Extinction coefficient (cm ^-1 M ^-1)	Correction Factor (280 nm)
ReadiUse™ Preactivated APC	651	660	730000	0.195
ReadiUse™ Preactivated PerCP	477	678	406000	0.22

References

View all 96 references: Citation Explorer

Imaging Flow Cytometry for Phylogenetic and MorphologicallyBased Functional Group Clustering of a Natural Phytoplankton Community over 1 Year in an Urban Pond.
Authors: Dunker, Susanne
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)

In Vivo Photodynamic Therapy With a Lipophilic Zinc(II) Phthalocyanine Inhibits Colorectal Cancer and Induces a Th1/CD8 Antitumor Immune Response.
Authors: Chiarante, Nicolás and Duhalde Vega, Maite and Valli, Federico and Zotta, Elsa and Daghero, Hellen and Basika, Tatiana and Bollati-Fogolin, Mariela and García Vior, María C and Marino, Julieta and Roguin, Leonor P
Journal: Lasers in surgery and medicine (2020)

Purification of Live Stem-Cell-Derived Islet Lineage Intermediates.
Authors: Rasouli, Niloofar and Melton, Douglas A and Alvarez-Dominguez, Juan R
Journal: Current protocols in stem cell biology (2020): e111

Application Settings versus Fixed Photomultiplier Tube Voltages for Optimal Cytometer Stability over Time, and Effect on Reusing a Compensation Matrix.
Authors: Andersen, Morten Nørgaard and Skovbo, Anni and Petersen, Charlotte Christie and Hokland, Marianne
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)

Improved Flow Cytometric Light Scatter Detection of Submicron-Sized Particles by Reduction of Optical Background Signals.
Authors: Arkesteijn, Ger J A and Lozano-Andrés, Estefanía and Libregts, Sten F W M and Wauben, Marca H M
Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology (2020)

Page updated on November 20, 2024

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Catalog Number	2560
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Fax	1-800-609-2943
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Physical properties

Solvent

Water

Spectral properties

Correction Factor (280 nm)	0.175
Extinction coefficient (cm ^-1 M ^-1)	1960000
Excitation (nm)	565
Emission (nm)	574
Quantum yield	0.82

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
UNSPSC	12171501