Serum Proteomics

Serum & Plasma proteomics services can identify low abundant disease-specific serum/plasma protein markers for therapeutics and diagnostics.

We have developed a Serum and Plasma 2D DIGE proteomics protocol by which thousands of low-abundance serum proteins can be resolved and quantitatively analyzed WITHOUT immunoaffinity depletion of the serum abundant proteins. This technology allows for a more sensitive detection and accurate quantification of low-abundance serum/plasma proteins. For details please view see our publication in Genetic Engineering & Technology News. Such platform has been successfully applied in discovering disease- or treatment-specific serum/plasma biomarkers.

Why does serum/plasma proteomics work best with 2D DIGE but not LC-MS/MS?

Because 2D DIGE can generate much better serum proteomics data at a lower cost. Analysis of serum and plasma proteome represents a challenge due to the wide range of protein concentrations and high structural complexity of the constituent proteins in these samples. In serum and plasma, the quantities of high- and some low-abundance proteins span over 10 orders of magnitude. Immunodepletion has been used to enhance the detection of low abundance plasma proteins. However, researches have discovered that, even with immunodepletion, low abundance proteins still remain undetectable using LC-MS/MS platform. Furthermore, immunodepletion process will: (i) remove proteins associated with the abudant proteins, (ii) cause protein degradations, and (iii) introduce variations, thus compromising the data quality.

Our serum/plasma proteomics services:

• A. Improved serum 2D DIGE without immuno-depletion
• B. Improved serum 2D DIGE combined with immuno-depletion of abundant proteins

For gel layout, please view the study designs based on the total number of samples.

A. Improved Serum 2D DIGE Without Immuno-depletion

Example 1: Human Serum Sample

The left, middle and right image shows the human serum proteome using standard protocol, Applied Biomics (ABO) improved protocol with pH3-10, and pH4-7, respectively. The ABO protocol clearly reveals much more low abundant serum proteins than the standard protocol.

Standard 2D Protocol (pH 3-10)

ABO pH 3-10 Protocol

ABO pH 4-7 Protocol

Example 2: Preclinical Trial Mouse Serum Sample

Four pairs of mouse serum samples were collected before (Control) and after 4 different treatments (Treated). 2D DIGE was performed using Applied Biomics’ improved serum protocol. The 4 Control (red) /Treated (green) overlay images showed the differences between Control and Treated samples. The circled spots represent very low abundant proteins which are up-regulated due to drug treatment. The 3-D view of the same spot is displayed in the right panel. The data shows that ABO protocol significantly improves the detection of low abundant proteins in the serum samples.

B. Improved Serum 2D DIGE Combined With Immuno-depletion Of Abundant Proteins

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

Protein dynamic range: 10 magnitude of difference in abundance

We offer the following serum abundant protein immuno-depletion services:

1. Human Serum: Albumin/IgG depletion
2. Human Serum: IgY-12 depletion (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).
3. Mouse Serum: Albumin/IgG depletion
4. Rat Serum: Albumin/IgG depletion
5. Monkey Serum: Albumin/IgG depletion

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

Plasma Without Immuno-Depletion

Plasma With Immuno-Depletion