Valosin-containing protein, Abcam, ab109240

Our team has carried out deep research in the published scientific literature to identify papers and reports using antibodies directed against human Valosin-containing protein (VCP) and followed up by extraction and evaluation of the available data to assess its performance according to pAbmAbs rating criteria.

YCharOS is here conducting a large head-to-head study comparing 16 commercial research antibodies against human VCP.

The collection of antibodies was tested using U2OS WT cells and cells with siRNA mediated knock-down of VCP (VCP KD) in the in the applications WB, IP and IF according to YCharOS standard protocols (see reference).

WB was performed using lysates (10 µg protein) from WT and KD cells. IP was performed using WT cell lysates, and immunoprecipitated fraction was analysed by WB using anti-VCP MA3-004 from Thermo Fisher Scientific. IF staining was performed using WT and KD cells.

WB with ab109240 (antibody dilution 1:50.000) produced a single intense band of approximately 90 kDa in lysate from WT U2OS cells, with no corresponding band detected in KD lysate.

IP (using 2 µg antibody) yielded a distinct band at 90 kDa, with higher intensity than the corresponding band in the input extract (4% of starting material). A clear, albeit reduced, band was also observed in the unbound fraction (4% of total material), indicating that the antigen was not completely depleted. In addition, a 50 kDa band was detected in the immunoprecipitated fraction, likely due to cross-reactivity of the WB secondary antibody with the Ig heavy chain. A weak background band (>100 kDa) band was also observed.

In IF (antibody dilution 1:500), a clear intense staining was observed in WT cells with no background staining in KD cells.

We highly encourage other scientists to share their observations.