I'm working with both Native PAGE gels and SDS under non reducing (SDS-NR) conditions gels.
My understanding is that Native PAGE would resolve "native-like" proteins, and the SDS-NR bands would include: "native-like" + non-covalently bound aggregates + non native monomers. Thus, the SDS-NR band of the same sample, should be greater than or equal to the native like one.
However, I'm seeing that the SDS-NR band after my sample treatment completely disappears, whereas the native band it is reduced (compared to the control).. but does not disappear.
What am I doing wrong? I've already done this gels 4 times, with two different batches of samples :(
Thanks in advance!!
Mariana
PS. in the image C stands for control and H for heat-treated sample.
Non-reducing SDS denatures proteins and should produce bands for the individual monomeric proteins and disulfide-linked complexes. Heating the SDS samples usually helps to improve the denaturation and may result in fewer bands than in unheated samples (although it sometimes leads to aggregation). Therefore, you should expect fewer bands in the NR-SDS lanes than in the native lanes if there are noncovalent complexes and/or aggregates in the samples. In other words, your result is exactly what would be expected.
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