Glucagon receptor, Atlas Antibodies AB, HPA071228

We here review a publication from Novo Nordisk and University of Copenhagen providing a head-to-head comparison of all commercially available antibodies against the glucagon receptor (GCGR) (Bomholt et al., Communications Biology, 2022).
The authors systematically evaluated 12 commercial antibodies (Table S1) in HEK293 cells transfected with mouse or human cMyc-tagged GCGR cDNAs and studied liver sections from Gcgr+/+ and Gcgr-/- mice. Autoradiography, RNA-sequencing, and single cell RNA-sequencing were used as antibody independent approaches to support the findings obtained with IHC.
Transfected cells were permeabilized with methanol.
All GCGR antibodies, including this one from Atlas Antibodies, gave overlapping staining with the cMyc antibody, demonstrating the ability to specifically recognize recombinant GCGR in transfected cells.
No staining was seen in control transfected cells or cells without primary antibodies (Fig. 1).
The antibodies were also checked for their ability to detect membrane bound GCGR by staining unpermeabilized HEK293 cells transfected with human GCGR and fixed in 4% PFA.
Atlas Antibodies HPA071228 (Antibody 10 in the paper) also showed positive staining of membrane bound GCGR (Fig. S1)
The antibodies were subsequently evaluated using formalin-fixed and paraffin-embedded liver sections from female Gcgr+/+ and Gcgr-/- mice. However, the Atlas antibody showed no staining (Fig. 2a)
However, HPA071228 showed some signal in IHC on paraffin embedded human liver tissue from patients with NASH (Fig. 2S).
HPA071228 was further tested in Western blotting using transfected HEK293 cells as samples. Mock transfected cells were used as negative controls. HPA071228 detected a band corresponding to the predicted size of GCGR (55 kDa) but also bands of higher molecular weight, likely unspecific (Fig. 2b)
In conclusion, HPA071228 receives a 4 star rating in immunostaining based on its ability to specifically recognize GCGR in transfected cells and its positive signal in the human NASH samples, and a 2 star rating in Western blotting based the presence of high intensity unspecific bands.
We highly encourage the scientific community to share their experiences.