Hi. There are few alternatives to SDS PAGE. For example you can do ELISA, Flow cytometry or even PCR (to detect mRNA of your target protein). However, "inability to get any bands" is a vague field in itself, because detection of proteins in SDS PAGE is influenced by many factors including but not limited to the expression level of target protein in the cell type used, quality of sample preparation, amount of protein loaded (in most cases 30 ug is enough), gel concentration and other conditions of gel electrophoresis, concentration and quality of the primary and secondary antibody used, ECL solutions used and finally the machine used and resolutions selected for detection as well.

Therefore, before you jump into another technique first make sure about the expression level of protein in your cell type and if you think most conditions that I mentioned before are optimized, then the problem would lie either in the concentration of antibodies used or ECL solutions.

Hi there,

The issue is no staining at all or no specific band appearing when expressing your fav' protein?

Hi Chandini Inumala, you can improve the visualization of your bands on your gel staining with silver, this is a very sensitive method. Also, the quantification of the amount of proteins in each sample could be important. If you are looking for Western blot approach, you can check your transfer using Red Ponceau staining, is easy to use and very informative.

Hello Chandini Inumala

Visualization of protein bands is carried out by incubating the gel with a staining solution. You may continue using SDS PAGE to see the bands of proteins, but you could try changing the protein stain.

The most commonly used protein stain is Coomassie Blue staining, which is based on the binding of Coomassie Brilliant Blue and it binds non-specifically to virtually all proteins. Coomassie Blue staining is approximately 50-fold less sensitive than silver staining, however due to its simplicity binding Coomassie Blue is preferred.

Alternatively, you could use silver staining which is a very popular method for protein detection, allowing for the detection of nanogram quantities of proteins, and is able to detect 2–5 ng protein per band on a gel. However, an important consideration on the silver staining method is the fact that they are unable to detect all proteins, particularly glycoproteins and proteins with large modified groups attached to their side-chains. I recommend that you perform silver staining if you haven’t.

I am attaching the Protein Stains Handbook which could be helpful. You could choose the stain that would be most suitable for your needs.

http://wolfson.huji.ac.il/purification/PDF/PAGE_SDS/Protein_Staining/GBIOSC_ProteinStainHandbook.pdf

Best.

Malcolm Nobre Thank you.. i will go through it.

Yes, there are other alternatives to SDS PAGE for analyzing protein bands. These include isoelectric focusing (IEF), native PAGE, capillary electrophoresis (CE), gel filtration chromatography, and western blotting.

Bandla Ramesh Thank you.. I will look into it

Yes, there are other alternatives to SDS PAGE for viewing protein bands. These include native PAGE, two-dimensional PAGE, Western blotting, and non-denaturing PAGE.

Dominique Liger Smear path along the wells is visible.. But not proper bands..

Hi again,

So the migration occurs but resolution is very bad. The issue is either the gel composition or the sample composition... Or both... But not the staining as you see the smear... What about the behavior of MW markers?