What precautions should be taken when working with SDS PAGE?

Answer 1

(1) Use high quality acrylamide and bis.

(2)The high electrical power used in gel electrophoresis is very dangerous as such one should never disconnect the electrodes before first turning off the power supply.

3. The stacking gel length should be 1 cm from the well bottom to the top of the separating gel for proper stacking of the protein sample.

4. Band resolution could be improved by doubling the salt concentration in stacking and separating gels, but the gel must be run at lower voltages.

5. To avoid edge effects, add 1x sample buffer to unused wells.

6. If electrophoresis is carried out at low temperatures use lithium dodecyl sulfate (LiDS) instead of SDS. LiDS does not precipitate at low temperatures.

7. During protein sample treatment the sample should be mixed by vortexing before and after the heating step for best resolution.

8. Add 0.1mM thioglycolic acid to upper gel buffer if proteins will be subjected to sequence analysis.

9. When doing protein renaturation or sequencing applications, leave gel for at least 5 hours post-polymerization to allow the ammonium persulfate and the TEMED to react with the gel components which reduces their chance of reacting with the amino-terminal end of the peptide.

Answer 2

When working with SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis), it is important to wear appropriate personal protective equipment, such as gloves and lab coats. Additionally, ensure proper disposal of chemicals and avoid inhalation of toxic fumes by working in a well-ventilated area. Lastly, handle the gel and running buffer carefully to prevent spills and electric shock hazards.