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ReadiUse™ Preactivated PE-iFluor® 647 Tandem

Our preactivated PE-iFluor® 647 Tandem was premodified with our Buccutite™ FOL (provided). Your antibody (or other proteins) is modified with our Buccutite™ MTA (provided as free sample) to give MTA-modified protein (such as antibody). The MTA-modified protein readily reacts with FOL-modified PE-iFluor® 647 Tandem (provided) to give the desired PE-iFluor® 647 Tandem-antibody conjugate in much higher yield than the SMCC chemistry. In addition our preactivated PE-iFluor® 647 Tandem reacts with MTA-modified biopolymers at much lower concentrations than the SMCC chemistry.
Our preactivated PE-iFluor® 647 Tandem was premodified with our Buccutite™ FOL (provided). Your antibody (or other proteins) is modified with our Buccutite™ MTA (provided as free sample) to give MTA-modified protein (such as antibody). The MTA-modified protein readily reacts with FOL-modified PE-iFluor® 647 Tandem (provided) to give the desired PE-iFluor® 647 Tandem-antibody conjugate in much higher yield than the SMCC chemistry. In addition our preactivated PE-iFluor® 647 Tandem reacts with MTA-modified biopolymers at much lower concentrations than the SMCC chemistry.
Ordering information
Price ()
Catalog Number2577
Unit Size
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Additional ordering information
Telephone1-408-733-1055
Fax1-408-733-1304
Emailsales@aatbio.com
InternationalSee distributors
ShippingStandard overnight for United States, inquire for international
Physical properties
Molecular weight~240000
Spectral properties
Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)569
Emission (nm)666
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12171501
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OverviewpdfSDSpdfProtocol


Molecular weight
~240000
Extinction coefficient (cm -1 M -1)
1960000
Excitation (nm)
569
Emission (nm)
666
R-Phycoerythrin (PE) is isolated from red algae. Its primary absorption peak is at 565 nm with secondary peaks at 496 and 545 nm. Our RPE-iFluor® 647 Tandem is an excellent replacement for RPE-Cy5® and RPE-Alexa Fluor® 647 Tandem since they have almost identical spectra. ReadiUse™ Preactivated PE-iFluor® 647 Tandem is an activated PE protein, and can be easily conjugated to antibodies with much higher conjugation yield than the conventional PE.

Components


A: ReadiUse™ Preactivated PE-iFluor™ 647 Tandem1 vial (1 mg)
B: Buccutite™ MTA1 vial (100 µg)
C: Spin Desalting ColumnNot Included

Example protocol


AT A GLANCE

Important      PE-iFluor™ 647 Tandem 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-iFluor™ 647 Tandem (provided) to give the desired PE-iFluor™ 647 Tandem antibody conjugate.

SAMPLE EXPERIMENTAL PROTOCOL

Preparation of pre-activated Antibody with Buccutite™ MTA
  1. 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.
  2. Prepare target antibody (Ab) in pH = 8.5 - 9.0 buffer at a concentration above 1 mg/ml.
  3. Add the MTA to Ab solution at the ratio of 8 - 10 µg MTA/100 µg Ab.
  4. Mix well and react at room temperature for 60 minutes, rotating during the reaction.
  5. Purify the reaction mixture with a desalting column to remove any unreacted MTA. Exchange the buffer to PBS or another buffer of your choice.
  6. Collect the MTA-activated Ab. Estimate the concentration by 70% yield of the original starting amount. 

Conjugate with Pre-activated PE-iFluor™ 647 Tandem
  1. Reconstitute pre-activated PE-iFluor™ 647 Tandem in 100 µL ddH2O to 10 mg/mL.
    Note     Reconstituted pre-activated PE-iFluor™ 647 Tandem is not stable and can not be stored for more than one month.
  2. Add pre-activated PE-iFluor™ 647 Tandem directly to MTA-activated target Ab solution at the ratio of 300 µg PE-iFluor™ 647 Tandem/100 µg MTA-activated Ab.
  3. Rotate the mixture for 1 - 2 hours at room temperature.
  4. The Ab/PE-iFluor™ 647 Tandem 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-iFluor™ 647 Tandem can be stored at 4 °C for two months.
  5. Optional: Ab/PE-iFluor™ 647 Tandem can be further purified through size exclusion chromatography to get better performance. 

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)569
Emission (nm)666

Product family


NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)
ReadiUse™ Preactivated PE-iFluor® 750 Tandem566778-
ReadiUse™ Preactivated PE-iFluor® 594 Tandem5666061960000
ReadiUse™ Preactivated PE-iFluor® 700 Tandem5667081960000
ReadiUse™ Preactivated PE-iFluor® 660 Tandem5666951960000

References


View all 46 references: Citation Explorer
Chromophore attachment to phycobiliprotein beta-subunits: phycocyanobilin:cysteine-beta84 phycobiliprotein lyase activity of CpeS-like protein from Anabaena Sp. PCC7120
Authors: Zhao KH, Su P, Li J, Tu JM, Zhou M, Bubenzer C, Scheer H.
Journal: J Biol Chem (2006): 8573
Excitation energy transfer from phycobiliprotein to chlorophyll d in intact cells of Acaryochloris marina studied by time- and wavelength-resolved fluorescence spectroscopy
Authors: Petrasek Z, Schmitt FJ, Theiss C, Huyer J, Chen M, Larkum A, Eichler HJ, Kemnitz K, Eckert HJ.
Journal: Photochem Photobiol Sci (2005): 1016
Single-molecule spectroscopy selectively probes donor and acceptor chromophores in the phycobiliprotein allophycocyanin
Authors: Loos D, Cotlet M, De Schryver F, Habuchi S, Hofkens J.
Journal: Biophys J (2004): 2598
Isolation and characterisation of phycobiliprotein rich mutant of cyanobacterium Synechocystis sp
Authors: Prasanna R, Dhar DW, Dominic TK, Tiwari ON, Singh PK.
Journal: Acta Biol Hung (2003): 113
Evaluation of Tolypothrix germplasm for phycobiliprotein content
Authors: Prasanna R, Prasanna BM, Mohammadi SA, Singh PK.
Journal: Folia Microbiol (Praha) (2003): 59
Co-ordinated expression of phycobiliprotein operons in the chromatically adapting cyanobacterium Calothrix PCC 7601: a role for RcaD and RcaG
Authors: Noubir S, Luque I, Ochoa de Alda JA, Perewoska I, T and eau de Marsac N, Cobley JG, Houmard J.
Journal: Mol Microbiol (2002): 749
Phycobiliprotein genes of the marine photosynthetic prokaryote Prochlorococcus: evidence for rapid evolution of genetic heterogeneity
Authors: Ting CS, Rocap G, King J, Chisholm SW.
Journal: Microbiology (2001): 3171
Phycobiliprotein-Fab conjugates as probes for single particle fluorescence imaging
Authors: Triantafilou K, Triantafilou M, Wilson KM.
Journal: Cytometry (2000): 226
Novel activity of a phycobiliprotein lyase: both the attachment of phycocyanobilin and the isomerization to phycoviolobilin are catalyzed by the proteins PecE and PecF encoded by the phycoerythrocyanin operon
Authors: Zhao KH, Deng MG, Zheng M, Zhou M, Parbel A, Storf M, Meyer M, Strohmann B, Scheer H.
Journal: FEBS Lett (2000): 9
Phycobiliprotein and fluorescence immunological assay
Authors: Wu P., undefined
Journal: Sheng Li Ke Xue Jin Zhan (2000): 82