Specific: ca. 110 kDa Non-specific: ca. 65, ca. 30 kDa
Human hematopoietic cell lines, e.g. Jurkat and THP-1 cells and B16 cells transfected with NCKAP1L NB!! These positive controls did not always display a specific NCKAP1L band!! The band pattern of B16 with or without NCKAP1L transfection was also similar.
B16 cells not transfected with NCKAP1L
DISCOUNT CODE: 100%ABR-E7BVB
There are concerns whether this antibody is specific to NCKAP1L, because it also gives a "specific" band around the expected height in non-hematopoietic cells. This unexpected, since NCKAP1L expression is as mentioned in the product sheet: "Expressed only in cells of hematopoietic origin". The suggested positive control, 293T cells, should therefore also not express NCKAP1L and would be more likely to be a negative control. Possibly there is cross reactivity with Nap1, which is expressed in adherent cells.
After looking over your protocol it may help to reduce the concentration of the antibody that you are using. We recommend using a 1 ug/ml concentration which would be a 1/1000 dilution. Additionally, adding more of the blocking agent to the primary antibody for the incubation can also help reduce background.
Abcam user community
Submitted Jul 21 2011
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