Md Arif Hussain , I guess SDS amount or some surfactant amount are not enough. I have not handled prolamins, but I found prolamin had poor solubility and was soluble in around 50% ethanol by Google search. Try modification your sample preparation method, adding extra SDS (to 1-2%), or glycerol (polyol is sometimes effective as surfactant), etc? Some people was succeeded in SDS-PAGE of prolamin and glutelin.

https://www.researchgate.net/publication/314017944_Isolation_and_characterization_of_gluten_protein_types_from_wheat_rye_barley_and_oats_for_use_as_reference_materials/figures. See Fig.4.

Md Arif Hussain , I guess SDS amount or some surfactant amount are not enough. I have not handled prolamins, but I found prolamin had poor solubility and was soluble in around 50% ethanol by Google search. Try modification your sample preparation method, adding extra SDS (to 1-2%), or glycerol (polyol is sometimes effective as surfactant), etc? Some people was succeeded in SDS-PAGE of prolamin and glutelin.

https://www.researchgate.net/publication/314017944_Isolation_and_characterization_of_gluten_protein_types_from_wheat_rye_barley_and_oats_for_use_as_reference_materials/figures. See Fig.4.

@ Kou Hayakawa Thank you for your valuable suggestionS.

Michitoshi Watanabe Thank you very much for your information. Can you suggest how much should I add / amount should I apply for glycerol . I tried another way , open the lid of sample for 4/5 hrs (-5) , but can’t find expected result .

Md Arif Hussain If you can, see Journal of Cereal Science Volume 56, Issue 3, November 2012, Pages 726-732, ref.31 in the article I introduced above. They use the buffer including 2% SDS to extract prolamin, i.e. they collect and use the soluble fraction by 2% SDS for SDS-PAGE. I think you will be able to resolve this issue, how about?

And add 20%(w/v) SDS solution (water is better for solvent) to desired concentration when you want to add extra SDS. But sample density become lower, add extra glycerol or sucrose to keep sample density. Please search required concentration of glycerol or sucrose for SDS-PAGE samples. Good Luck!!

@Michitoshi Watanabe

Got this . And I will try this . Thanks a lot for your comments .

I agree about the addition of glycerol. I find it very much improves loading for many proteins, especially less hydrophilic ones.

Md Arif Hussain

Please see the following link:

30 µL aliquots from whole wheat flour [5 mg] extracted with 250 µl of sample buffer

Article Electrophoretic Characteristics of Gluten Proteins as Influe...

@Dr Anju Kaushal

Thank you very much for your information .

Michitoshi Watanabe Kou Hayakawa

Md Arif Hussain

great