I am currently trying to detect eNOS monomer/dimer on western blot (a difficult endeavour...). Most of the articles in literature stress the importance of low-temperature elektrophoresis.
I homogenize with a mixer in 1xRIPA buffer. I do not heat the sample during incubation with laemmli buffer (1:1). When I do not heat, the sample becomes incredibly viscous and cannot be pipetted in the well of the gel. It simply sticks on the pipette tip and does not 'fall' in the well.
I was wondering if anyone has tips to circumvent this problem and can load a decent sample without boiling Laemmli.
When I do boil the sample with the Laemmli, everything is just fine.
Thanks in advance!