Huntingtin, Abcam, ab45169

Our team has carried out deep research in the published scientific literature to identify papers and reports using antibodies directed against human Huntingtin 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 20 commercial research antibodies against human Huntingtin encoded by HTT.

The collection of antibodies was tested using DMS35 WT and HTT KO cells in the applications WB (WT and KO cell lysates/30 µg protein), IP (WT cell lysate) and IF (WT and KO cells) according to YCharOS standard protocols (see reference). Furthermore, was this antibody evaluated in HEK 293T and HAP1in the application WB (WT and KO cell lysates/30 µg protein).

In WB ab45169 detected a double band of appr. 350 kDa in WT cell lysate with a faint corresponding band in KO cell lysate at 1:5000 ab dilution; several faint bands of lower molecular weight were observed in both genotypes. At 1:500 dilution ab45169 detected a double band at appr. 279 kDa in WT from DMS53, HEK293T, and HAP1 cells, with the signal strongly reduced or absent in the corresponding KO lysate. Faint bands of lower molecular weight were observed in all genotypes across cell lines. A double band corresponding to the target protein was detected in the IP sample following immunoprecipitation using 2 μg antibody. The signal appears stronger than in the input lane (4% of starting material), consistent with enrichment of the target protein during IP. A band was still present in the immunodepleted (unbound) fraction (4%), indicating that the antibody does not fully deplete the antigen from the lysate. In IF (1:500 Ab dilution), clearer staining was observed in WT cells, with minimal background signal in KO cells.

We highly encourage other scientists to share their observations.