Here is an example of Native PAGE protocol for acidic protein :

http://www.assay-protocol.com/molecular-biology/electrophoresis/native-page.html

Dear Subhayu Chakraborty

Would you like to perform Native PAGE or non reducing PAGE? or both? since non reducing could be even denaturing, since the S-S bonds are not destroed by SDS-page.

Non reducing but denaturing can be performed just avoid to add the reducing component in the laemil buffer used for sample load.

Reducing vd non reducing in SDS-page is used routinelly with purified monoclonal antibodies.

In this case Whith a so high MW you have to run a 3-8% tris-acetate gel, otherwise the protein will not enter in the gel.

Native SDS is little bit different and more complex

in this case i sugget you to give a look to:

https://www.thermofisher.com/it/en/home/life-science/protein-biology/protein-gel-electrophoresis/protein-gels/specialized-protein-gels/nativepage-bis-tris-gels.html

good luck

Manuele

To perform non-reducing SDS-PAGE for a native protein, prepare the sample without adding reducing agents like DTT or β-mercaptoethanol to the sample buffer, as these disrupt disulfide bonds. Use standard Laemmli buffer (with SDS to denature the protein) but omit the reducing agent. Load the protein samples mixed with this buffer onto a polyacrylamide gel (e.g., 10–12% depending on protein size). Run the gel under standard conditions (e.g., 80V for stacking, 120–130V for resolving) using SDS-PAGE running buffer. The absence of reducing agents preserves the protein's disulfide bonds, allowing you to analyze its native oligomeric or disulfide-linked state. After electrophoresis, stain the gel (e.g., Coomassie Blue) and proceed with visualization. Ensure fresh preparation of buffers to avoid contamination or oxidation artifacts.

Chinmay Hazare's answer is comprehensive. Run your sample using a loading buffer without a reducing agent, such as DTT or β-ME, to preserve disulfide bonds. The native maintains the protein structures without denaturants.