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ReadiUse™ Preactivated PE-Texas Red Maleimide

Flow cytometry analysis of whole blood stained with PE-Texas Red anti-human CD4 *SK3* conjugate. The fluorescence signal was monitored using an Aurora flow cytometer in the PE-Texas Red specific B6-A channel.
Flow cytometry analysis of whole blood stained with PE-Texas Red anti-human CD4 *SK3* conjugate. The fluorescence signal was monitored using an Aurora flow cytometer in the PE-Texas Red specific B6-A channel.
Ordering information
Price ()
Catalog Number2716
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 weightN/A
SolventWater
Spectral properties
Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)567
Emission (nm)615
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
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OverviewpdfSDSpdfProtocol


Molecular weight
N/A
Extinction coefficient (cm -1 M -1)
1960000
Excitation (nm)
567
Emission (nm)
615
PE-Texas Red is a popular tandem color used in flow cytometry. Its primary absorption peak is at 565 nm with emission peak at 600 nm. AAT Bioquest offers this preactivated PE-Texas Red maleimide to facilitate the PE-Texas Red 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-Texas Red maleimide is ready to conjugate the reduced antibodies and other thiol-containing biomolecules.

Example protocol


SAMPLE EXPERIMENTAL PROTOCOL

Reduction of IgG
  1. 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.
  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 temp for 30 minutes without additional mixing (to minimize reoxidation of cysteines to cystines).
  3. 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)
  4. 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-Texas Red Maleimide
  1. Reconstitute ReadiUse™ Preactivated PE-Texas Red Maleimide in 100 µL ddH2O to 10 mg/mL. Note: Reconstituted ReadiUse™ Preactivated PE-Texas Red Maleimide are stable at 4 °C for one week, please kept it from light.
  2. Add reduced antibody to pre-activated PE-Texas Red directly to at the ratio of 250-300 µg PE-Texas Red /100 µg reduced antibody.
  3. Rotate the mixture for 60-120 mins at room temperature.
  4. After 60 minutes, block the free sulfhydryls on the antibody.
  5. 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
  1. Antibody/PE-Texas Red conjugate could be further purified through size exclusion chromatography to get best performance. Note: The Antibody/PE-Texas Red conjugate solution is recommended to be stored at 2~8 °C and kept from light. 

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Extinction coefficient (cm -1 M -1)1960000
Excitation (nm)567
Emission (nm)615

Product family


NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)
ReadiUse™ Preactivated PE-Texas Red Tandem5676151960000

References


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Flow cytometry can diagnose classical hodgkin lymphoma in lymph nodes with high sensitivity and specificity.
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Journal: American journal of clinical pathology (2009): 322-32
Optimization of three- and four-color multiparameter DNA analysis in lymphoma specimens.
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Conjugation of fluorochromes to monoclonal antibodies.
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Journal: Cytometry (1996): 214-25
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Journal: Cytometry (1991): 570-8
Concomitant delineation of surface Ig, B-cell differentiation antigens, and HLADR on lymphoid proliferations using three-color immunocytometry.
Authors: Segal, G H and Edinger, M G and Owen, M and McNealis, M and Lopez, P and Perkins, A and Linden, M D and Fishleder, A J and Stoler, M H and Tubbs, R R
Journal: Cytometry (1991): 350-9