1. How much protein should I analyse?
We recommend analysing a 50ug whole cell lysate (total cellular
protein) or a 25ug nuclear extract per lane.
2. What percentage acrylamide gel should I use to
resolve the proteins?
<13 kDa 15% 14 - 50 kDa 12% 50 - 100 kDa 10% > 100 kDa
3. What membrane should I use?
We recommend using nitrocellulose. Although PVDF and other
similar membranes can be used, they can sometimes lead to increased
background particularly with goat polyclonal
4. What blocking buffer should I use?
We recommend using 1X TBS, 5% milk, 0.05% Tween-20. Blocking
should be conducted for 30-60 minutes at room temperature or
overnight at 4o. Please note that Tween should be omitted from the
buffer if incubating overnight.
5. What dilution of primary antibody should I
Check the datasheet of the individual antibody as this usually
notes starting dilutions. If there is not specific information on
the datasheet then we suggest that you perform a serial titration
against relevant positive and negative controls.