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Life Science > Proteomics > Sample Preparation > Protein Depletion > Seppro® Protein Depletion > Human IgY14 and SuperMix Columns
Protein Depletion

Human IgY14 and SuperMix Columns

Immunoaffinity partitioning of highly-abundant proteins (HAP) has proven to be one of the most effective approaches for overcoming the wide dynamic range of plasma protein concentration, as well as enabling the detection of low-abundant proteins (LAP). Avian polyclonal IgY (Immunoglobulin Yolk) antibodies have unique and advantageous features that allow highly-specific partitioning of protein mixtures. Sigma® offers a novel Seppro® IgY14 columns system, which takes you deeper than ever into the plasma proteome. IgY14 columns remove the top 14 HAP.

Proteins Partitioned by IgY-14 Microbeads:

  • HSA
  • IgG
  • Fibrinogen
  • Transferrin
  • IgA
  • IgM
  • Haptoglobin
  • alpha2-Macroglubulin
  • alpha1-Acid Glycoprotein
  • alpha1-Antitrypsin
  • Apo A-I HDL
  • Apo A-II HDL
  • Complement C3
  • LDL (ApoB)

Our IgY14 systems are available in the following formats (suitable for your volumes):


Product Name Column Use Human or Primate
plasma/serum
Product No.
IgY14 Spin Column Kit
Includes 2 spin columns and reagents.
Good for 200 samples.
100 times 20µl SEP010
IgY14 LC-2 Column Kit
Includes 1 HPLC column and reagents.
100 times 50µl SEP020
IgY14 LC-5 Column Kit
Includes 1 HPLC column and reagents.
100 times 100µl SEP030
IgY14 LC-10 Column Kit
Includes 1 HPLC column and reagents.
100 times 250µl SEP040


Human SuperMix System

Immunoaffinity fractionation of highly-abundant proteins (HAP) has proven to be one of the most effective approaches for overcoming the wide dynamic range of protein concentration, as well as enabling the detection of low abundant proteins (LAP). Avian polyclonal IgY (Immunoglobulin Yolk) antibodies have unique and advantageous features that allow for highly-specific fractionation of a complex protein mixture. The IgY14 column can effectively remove 14 HAP from plasma. The next level of abundant proteins, moderately abundant proteins (MAP), becomes an obstacle to access LAP, where the majority of biologically interesting and clinically important biomarkers reside. Therefore, isolation of MAP is a new challenge for effectively and accurately detecting and analyzing LAP.

To tackle this challenge, the SuperMix column was developed to separate MAP from LAP. The SuperMix System was developed by immunizing chickens with a flow-through fraction of IgY-12 or IgY14 column and constructing the column with affinity-purified IgY antibodies against the flow-through proteins of IgY-12 or IgY14.

SuperMix Fractionation Process

The Human SuperMix column is then used in conjunction with the IgY14 column to provide superior depletion.


Plasma Fractionation


SuperMix Results

As shown below, the SuperMix column was applied for fractionating the flow-through fraction of IgY14, which resulted in a bound/eluted fraction (designated MAP fraction) and the flow-through fraction (designated LAP fraction). Unfractionated and serial-fractionated samples using IgY14 and SuperMix columns were analyzed by SDS-PAGE and 2DE. Our data demonstrate that SuperMix columns specifically and reproducibly remove a group of proteins, which are the main abundant proteins in the flow-through fraction of IgY14.


1DE of IgY14

2DE of IgY14

 


Product Name Column Use Capacity Product No.
SuperMix LC-2 Column Kit
Includes single SuperMix LC-2 column and reagents.
100 times Flow-through from IgY14 LC-5 column SEP050 
SuperMix LC-5 Column Kit
Includes single SuperMix LC-5 column and reagents.
100 times Flow-through from IgY14 LC-10 column SEP060 

Recommended Set-Up:

Product Name Column Use Human or
Primate plasma
IgY14 LC-10/SuperMix LC-5
Columns are run in conjunction for intensive mass spectrometry and 2D-gel analysis
100 times 250µl
IgY14 LC-5/Supermix LC-2
Columns are run in conjunction for intensive mass spectrometry and 2D-gel analysis
100 times 100µl

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