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We offer the following services in serum 2D DIGE:

A. Improved Serum 2D DIGE without Immuno-depletion

B. Immuno-depletion of Serum Abundant Proteins


A. Improved Serum 2D DIGE Without Immuno-depletion

Applied Biomics has developed a protocol that significantly improves the performance of standard protocol. For details please click here to see our paper published on Genetic Engineering & Technology News. Please see below for examples using ABO (Applied Biomics) improved serum 2D DIGE protocol.


* Example 1: Human Serum Sample: significantly more protein spots can be profiled by ABO protocol.

Standard 2D protocol (pH3-10)
ABO pH3-10 protocol

ABO pH4-7 protocol

Proteomics Serum 2-D DIGE
Proteomics Serum 2-D DIGE
Proteomics Serum 2D DIGE

* Example 2: Analysis of Preclinical Trial Mouse Serum Before and After Drug Feeding Using ABO Protocol. Four paired mouse serum samples were collected before and after 4 different treatments, then each paired samples were analyzed by 2D DIGE using pH3-10 nonlinear IPG strip. The overlay images showed the difference between the serum samples collected before (in green color) and after (in red color) the drug feeding (18 hours). In the image overlay, the circled spots (in red) indicated a protein with increased abundance due to drug feeding. The 3-D view of the same spot is generated by DeCyder software.

Proteomics Serum 2D DIGE


B. Immuno-depletion Of Serum Abundant Proteins For 2D DIGE

The Challenge of Biomarker Discovery from Plasma/Serum

Transferrin: 76 kDa
Fibrinogen: 65 kDa (a), 51 kDa (g)
a1-antitrypsin: 56 kDa
IgG: 150 kDa
HSA: 67 kDa
IgA: 180 kDa
IgM: 750 kDa
Transferrin: 79 kDa
a2-Macroglobulin: 160 kDa
Haptoglobin: 43 kDa
alpha 1-acid glycoprotein: 23 kDa
APO A-I, APO A-II: 23 kDa

Proteomics Serum 2-D DIGE

Protein dynamic range: 10 magnitude of difference in amount
Procedure of Immuno-depletion of serum abundant proteins for 2D DIGE
1. Spin the serum and collect the clear supernatant
2. Dilute the serum with PBS containing the protease inhibitors
3. Equilibrate the immobilized IgY affinity columns (against serum abundant proteins)
4. Remove the abundant proteins by passing the diluted serum onto the affinity columns to
5. Collect the flow through
6. Concentrate the serum samples after immuno-depletion
7. Replace the buffer with 2D lysis buffer using 3 kDa MWCO spin column
8. Measure then adjust the protein concentration
9. Now samples are ready for 2D DIGE

We offer three kinds of serum abundant protein immuno-depletion services:

IgY-12 (for Human serum): against the 12 most abundant human plasma proteins: Human Serum Albumin, IgG, Fibrinogen, Transferrin, IgA, IgM, Haptoglobin, a2-Macroglobulin, a1-Acid Glycoprotein, a1-Antitrypsin and HDL (Apo A-I & Apo A-II).

IgY-M7 (for Mouse serum): against the 7 most abundant rodent (mouse) plasma proteins: Mouse Serum Albumin, IgG, Fibrinogen, Transferrin, a1-antitrypsin, Haptoglobin & IgM.

IgY-R7 (for Rat serum): against the 7 most abundant rodent (rat or mouse) plasma proteins: Rat Serum Albumin, IgG, Fibrinogen, Transferrin, a1-antitrypsin, Haptoglobin & IgM.

* Example: Human plasma with and without IgY-12 Immuno-depletion (pH4-7)

Plasma without immuno-depletion
Plasma after immuno-depletion
Proteomics Serum 2D DIGE Proteomics Serum 2D DIGE
Call our tech support at 510-887-0889 or send email to support@appliedbiomics.com with any technical questions.

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