Protein Phosphorylation

Purification Enrichment

Product no. Product Name Function Format Samples
Processed
Input Sample Size Output
Compatibility
P9740 PHOS-Select™ Iron Affinity Gel Agarose Fe+3-chelate phosphopeptide capture Bulk media 5 Captures 4 µM phosphopeptide maximum per mL packed media LC, MALDI-MS, ESI-MS


PHOS-Select™ Iron Affinity Gel (P9740)

  • High capacity (at least 4 micromoles of phosphopeptide per ml of resin)
  • Precharged with iron (III) as a stable chelate
  • Provides single-step phosphocompound purification
  • Supplied as an approximate 50% suspension in 50% glycerol buffered at pH 5.0
  • Supplied with technical bulletin providing detailed procedure for use

Workflow highlighting the use of the spin columns for digestion and selective enrichment of phosphopeptides from sample.

Workflow highlighting the use of the spin columns for digestion and selective enrichment of phosphopeptides from sample.

 

Binding comparison: Selected portions of HPLC chromatograms demonstrating enrichment of phosphopeptides for IMAC technologies.
Standard phosphopeptides representing the three most common sites of phosphorylation phosphoserine, phosphothreonine, and phosphotyrosine were used. The lyophilized solids were first dissolved in water and an approximately equimolar mixture of the peptides was formulated. Each phosphopeptide was added to the BSA digest at a weight ratio of ~1.7% to produce a total phosphopeptide content of ~5% by weight. Quantitation results are given below. Note that competitor A, B, and C technologies were biased in selecting Peptide 2, while our technology bound and eluted the peptides in approximately the same ratio as applied to the column.

HPLC Chromatograms comparing phosphopeptide enrichment by IMAC methods

 

Kit Recovery of phosphopeptide standards Specificity*
1 2 3 Total
Ours 59% 52% 74% 59% 50%
Competitor A 6% 19% 11% 13% 28%
Competitor B 39% 56% 17% 42% 28%
Competitor C 37% 65% 37% 46% 25%

 

Additional Products for Phosphoprotein Analysis

Immobilized Antibodies for the Isolation of Phosphopeptides  
Product Name Product No.
Monoclonal Anti-Phosphotyrosine-Agarose from mouse, Purified immunoglobulin, PBS suspension, clone PT-66 A1806
Monoclonal Anti-Phosphoserine-Agarose, Purified immunoglobulin, PBS suspension, clone PSR-45 A8076
   
Controls and Elution Reagents  
Product Name Product No.
Phosphotyrosine-BSA Phosphotyrosine Antibody Inhibitor, 2 mg per ml, Buffered aqueous solution, for blotting and ELISA P3967
Phosphoserine-BSA, Phosphoserine Antibody Inhibitor, 2 mg/ml, for blotting and ELISA P3717
Phosphothreonine-BSA, Phosphothreonine Antibody Inhibitor, 2 mg/ml, for blotting and ELISA P3842
   
Resins and Plates  
Product Name Product No.
Avidin-agarose from egg white, Aqueous glycerol suspension A9207
Streptavidin-Agarose from Streptomyces avidinii, Buffered aqueous suspension S1638
SigmaScreen Streptavidin 96-well High Capacity Coated Plates S6940
Anti-Mouse IgG (whole molecule)-Agarose from goat, IgG fraction of antiserum, Saline suspension A6531
EZview Red Protein A Affinity Gel P6486
EZview Red Protein G Affinity Gel E3403
Protein L-Agarose from Peptostreptococcus magnus, recombinant, expressed in Escherichia coli P3351

 

 Phosphatase Inhibitor Cocktails

 
Product No. P5726 P2850
  Tyrosine Protein Phosphatases and Acid and Alkaline Phosphatases Ser/Thr Protein Phosphatases and Alkaline Phosphatase L-Isozymes
(--)-p-Bromotetramisole  
Cantharidin  
Imidazole  
Microcystin LR  
Sodium molybdate  
Sodium orthovanadate  
Sodium tartrate  


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