Anti-HGS antibody

• IP

Supplier Data

Immunoprecipitation - Anti-HGS antibody (AB72053)

Detection of human HGS by western blot of immunoprecipitates.
Samples : Whole cell lysate (0.5 or 1.0 mg per IP reaction; 20% of IP loaded) from HeLa cells prepared using NETN lysis buffer.
Antibodies : ab72053 used for IP at 6 μg per reaction, lanes 1 and 2, lane 3 is control IgG. For blotting immunoprecipitated HGS, ab72053 was used at 0.4 μg/ml.

All lanes:

Immunoprecipitation - Anti-HGS antibody (ab72053)

Predicted band size: 86 kDa

false

Exposure time: 3s

• WB

Supplier Data

Western blot - Anti-HGS antibody (AB72053)

Lysates prepared using NETN lysis buffer.

All lanes:

Western blot - Anti-HGS antibody (ab72053) at 0.1 µg/mL

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 50 µg

Lane 2:

HeLa whole cell lysate at 15 µg

Lane 3:

HeLa whole cell lysate at 5 µg

Predicted band size: 86 kDa

false

Exposure time: 10s

• WB

Supplier Data

Western blot - Anti-HGS antibody (AB72053)

Lysates prepared using NETN lysis buffer.

All lanes:

Western blot - Anti-HGS antibody (ab72053) at 0.1 µg/mL

Lane 1:

Renca (Mouse renal adenocarcinoma cell line) whole cell lysate at 15 µg

Lane 2:

CT26.WT (Mouse renal colorectal carcinoma cell line) whole cell lysate at 15 µg

Lane 3:

4T1 (Mouse mammary gland cell line) whole cell lysate at 15 µg

Predicted band size: 86 kDa

false

Exposure time: 3min

• ICC/IF

CiteAb

Immunocytochemistry/ Immunofluorescence - Anti-HGS antibody (AB72053)

Immunocytochemistry-immunofluorescence using Anti-HGS antibody, ab72053. Publication image from Sánchez-Madrid, F. et al., 2016, Nat Commun, 27882925. Legend direct from paper.

ISG15 conjugation induces protein aggregation and degradation by lysosomes.(a) Confocal co-localization analysis of ISG15-GFP (green) and the MVB marker HRS (red). Right graphs : fluorescence intensity profiles of ISG15-GFP (green) and HRS (red) in the regions delineated by a white line. Nuclei were stained with DAPI. Scale bar, 10 µm. (b) Confocal co-localization analysis of ISG15-GFP (green) and p62 (red). Right graphs represent fluorescence intensity profiles of ISG15-GFP (green) and p62 (red) of the regions delineated by a white line. Nuclei were stained with DAPI. Scale bar, 10 µm. (c) Confocal microscopy analysis of HRS (green) and LAMP1 (red) co-localization in HEK293 cells co-transfected with ISGylation machinery and functional (ISG15WT) or mutated ISG15 (ISG15MUT). Nuclei were stained with DAPI. Scale bar, 10 µm. Co-localization area per cell was quantified by ImageJ (n=16). Each dot represents the percentage of co-localization area per cell surface and mean is indicated in red lines. (d) Western blot analysis of exogenous ISG15 in HEK293 cells transfected with ISG15-GFP. Cells and EVs (EXO) were blotted for GFP and CD81. (e) Western blot analysis of ISG15-GFP and GFP in 0.5% NP-40 soluble and insoluble cell fractions in HEK293 transfected with GFP or ISG15-GFP. Right graph : quantification of GFP and ISG15-GFP in the insoluble fraction respect to the soluble fraction in three independent experiments. (f) Western blot analysis of GFP and ISG15-GFP degradation kinetics in HEK293 cells transfected with GFP or ISG15-GFP and treated with cycloheximide to inhibit protein synthesis during the indicated times. Where specified, the medium was supplemented with the lysosome inhibitor Bafilomycin A1 (BAF) or the proteasome inhibitor MG132. A representative blot from two independent experiments is shown. Data from c,e : t-test *P-value<0.05 and ***P-value<0.0001.