What is an issue with using 2D-PAGE?
a. Hydrophobic proteins may not run as expected due to the hydrophobic surfaces.
b. Some proteins may not migrate through polyacrylamide and therefore not be represented on the gel.
c. Rare cellular proteins are hard to visualize with Coomassie blue protein stain.
d. All of the above are issues with 2D-PAGE

The correct answer is:

(d) All of the above are issues with 2D-PAGE.

Explanation:

2D-PAGE (Two-Dimensional Polyacrylamide Gel Electrophoresis) is a powerful technique for separating complex protein mixtures based on both isoelectric point (pI) and molecular weight. However, it has several limitations:

1. (a) Hydrophobic proteins may not run as expected due to the hydrophobic surfaces

   • Membrane proteins are often hydrophobic and tend to precipitate or aggregate instead of properly migrating through the gel.
   • This makes them difficult to resolve in 2D-PAGE.

2. (b) Some proteins may not migrate through polyacrylamide and therefore not be represented on the gel

   • Very large proteins or proteins with abnormal structures may not enter or migrate properly through the gel.
   • This can result in incomplete representation of the sample.

3. (c) Rare cellular proteins are hard to visualize with Coomassie blue protein stain

   • Coomassie blue has a low sensitivity, meaning that low-abundance proteins may not be detectable.
   • More sensitive stains (e.g., silver staining, SYPRO Ruby, or fluorescent dyes) are needed to visualize rare proteins.

Conclusion:

Since all the issues listed affect 2D-PAGE, the correct answer is:

✅ (d) All of the above are issues with 2D-PAGE.