Hi,

If you want to use gel based proteomic approach the best strategy is to use two dimensional electrophoresis (2DE). But comparison of two 2DE maps can be a difficult task. In order to avoid that one can use 2-D Fluorescence Difference Gel Electrophoresis (2-D DIGE) which  is a variant of two-dimensional gel electrophoresis.  This offers the possibility to include an internal standard so that all samples can easily be compared and accurately quantitated.

But gel-based proteomic strategies are rapidly being supplanted by methods that involve peptide separation via high efficiency nanocolumn liquid chromatography separation techniques directly linked to mass spectrometer. However some protein samples present complexity beyond separation capacity of a 1D LC techniques. The enhanced separation capacity, which is important for global expression studies, can be achieved by a technique known as MudPIT (Multidimensional protein expression technology), Link et al, Nature Biotechnollogy, 17: 676-682.

Having said all this, the technique of choice depends on the facilities one have access to.

as discussed above, I would say that 2D-DIGE is by far the best method to do these kind of analysis. 

I recently came across a nice review about proteomic tools for environmental microbiology. It is a good overview and the methods should be applicable for your research as well.

http://www.ncbi.nlm.nih.gov/pubmed/23894077

Thanks all for your suggestions. 

We thought Quantitative proteomics might give us the individual differentially regulated proteins, but due to the experiment cost it may not be feasible at this time point and same goes for 2D-DIGE as well. I could certainly use 2D electrophoresis.

I would recommend mass spectrometry-based proteomics discovery methods if you have decent equipment available. After the initial discovery phase, you might try selective reaction monitoring - if the (triple quad) equipment is available

You can use a new methodology based on gel-free approach (i.e. LC-MS-MS). It is less labor than 2D, but the cost is twice higher. In fact, this approach used a liquid chromatography separation coupled with a MS detector. You can obtain a better separation of protein, and so a higher number of identified protein.

If you going to use 2DE, make sure to use sensitive staining method. Silver staining or similar. With silver staining you might get higher background. You could use the  method called blue silver. It's very sensitive, cheap to make and no background.

There are a variety of technologies around that come to my mind

> protein arrays (you can test a couple of thousand interactions in a single experiment)

Protein arrays bypass the genomic biomarker discovery and validation stage and focuses straight away on biomarkers that are detectable in the blood stream. There have been many promising biomarkers been discovered in biopsies and tissues that never translated in biomarkers that are detectable in the blood stream. This was always due to dilution effects from and markers are not sufficiently expressed on the proteome level.

There are a number of diseases and applications such as predictive as well as efficacy biomarker for research and for clinical trial monitoring.

Cancer and Autoimmune Disease biomarker discovery

Measuring global immune response to drugs in Clinical Trials

Monitoring global immune response to viral, microbial and fungal infection

There are proper tried and tested workflows for protein arrays around including service labs that do all the work for you.. I have performed -50 of those studies.

Protein arrays can give you quantitative results for each of you samples and then you can perform statistical analysis on your results get a histogram as shown below.

Other technologies:

> MSD 

> mass spec 

Feel free to get in touch if you require more information.

Jens

http://www.biomarker-discovery.com/