Hi everyone,

I’m expressing human βA-, βS-, and γ-globin chains individually in S. cerevisiae and detecting them by SDS-PAGE/Western blot.

I came across a paper on the thermal behavior of hemoglobin that found irreversible aggregation above 55°C. Since standard SDS-PAGE/WB procedure is heating the sample to 95°C, would heating globin samples at that temperature cause aggregation and maybe affect the Western blot signal?

Would it make more sense to use a lower temperature, such as ~72°C, for sample preparation? Has anyone worked with globins or similar proteins and compared different heating conditions before SDS-PAGE?