Ataxin-3, Proteintech, 13505-1-AP

Our team has carried out deep research in the published scientific literature to identify papers and reports using antibodies directed against human Ataxin-3 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 seven commercial research antibodies against human Ataxin-3 encoded by ATXN3.

The collection of antibodies was tested using HEK293T WT and ATXN3 KO cells in WB (WT and KO cell lysates/30 µg protein). Furthermore antibody 13505-1-AP, was additionally tested in HEK293T, U2OS, NPC (WT and ATXN3 KO) and HAP1 WT. The antibodies were also tested in the applications IP (HEK293T WT cell lysate) and IF (U2OS WT and ATXN3 KO cells) according to YCharOS standard protocols (see reference).

In WB (1:1000 Ab dilution) 13505-1-AP detected a double band of appr. 42 kDa in WT cell lysate, with no band observed in KO cell lysate. A clear band of higher molecular weight was observed in both genotypes. Additionally, western blot of lysates of HEK293T, U2OS, NPC (WT and ATXN3 KO) and HAP1 WT was carried out. Antibody 13505-1-AP was used at 1:1000. A double band was observed in HEK293T, U2OS, NPC and HAP1 WT, with no corresponding band observed in KO cell lysates. A double band was also detected following IP (1 µg Ab used). This double band was marked more intense than the corresponding band in input extract (10 % of starting material), Furthermore, no band was observed in immunodepleted extract (10% of total unbound fraction) showing that the antibody does completely deplete the antigen from the cell lysate. In IF (1:500 ab dilution) WT cells had more intense staining compared to KO cells, which had background staining.

We highly encourage other scientists to share their observations.