Anti-Sumo 1 antibody [Y299] - ChIP Grade

• ICC/IF

PubMed

Immunocytochemistry/ Immunofluorescence - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Immunofluorescence analysis of ICP0-null mutant HSV-1 infected HepaRG cells, staining Sumo1 (green) with ab32058. An AlexaFluor®-conjugated goat anti-rabbit IgG was used as the seconday antibody.

Image from Cuchet-Lourenço D et al. PLoS Pathog. 2011 Jul;7(7):e1002123. Epub 2011 Jul 14. Fig 9.; doi:10.1371/journal.ppat.1002123; July 14 2011 PLoS Pathog 7(7): e1002123.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluor^® 488) at a dilution of 1/2000 was used as the secondary antibody.

Isoytype control : Rabbit monoclonal IgG (Black)

Unlabelled control : Cell without incubation with primary antibody and secondary antibody (Blue)

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labelling Sumo 1 with ab32058 at 1/20 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluorr^®488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in human bladder carcinoma tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/250. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1 : PBS in place of primary antibody.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

IHC of paraffin-embedded human lung carcinoma using anti-SUMO 1 (ab32058) diluted 1 : 250

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Immunofluorescent staining of HeLa (Human epithelial cell line from cervix adenocarcinoma) cells using anti-SUMO 1 (ab32058) diluted 1/250.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with 4% Paraformaldehyde and permeabilized with 0.1% Triton X-100. Samples were incubated with primary antibody at a dilution of 1/500. A goat anti rabbit IgG (Alexa Fluor® 488) (ab150077) was used as the secondary antibody at a dilution of 1/1000. ab195889 was used as a counterstain for primary antibody ab133645 at 1/200. DAPI was used as a nuclear counterstain and PBS as a negative control.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in human endometrium tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/250. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1 : PBS in place of primary antibody.

• ChIP

Unknown

ChIP - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Chromatin was prepared from SK-OV-3 (Human ovarian cancer cell line) cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25μg of chromatin, 5μg of ab32058 (red, and 20μl of Anti rabbit IgG sepharose beads. 5μg of rabbit normal IgG was added to the beads control (grey). The immunoprecipitated DNA was quantified by real time PCR (Sybr green approach).

ChIP was performed according to the literature (PMID : 23770046).

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in rat stomach tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/250. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1 : PBS in place of primary antibody.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 staining Sumo 1 in mouse kidney tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and antigen retrieval was by heat mediation in a EDTA buffer. Samples were incubated with primary antibody at a dilution of 1/250. A goat anti-rabbit IgG H&L (HRP) ab97051 was used as the secondary antibody at a dilution of 1/500.

Negative control 1 : PBS in place of primary antibody.

• IP

Unknown

Immunoprecipitation - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

ab32058 immunoprecipitating Sumo 1. 10μg of NIH/3T3 (Mouse embryonic fibroblast) cell lysate was incubated with primary antibody at a dilution of 1/20 and VeriBlot for IP Detection Reagent (HRP) (ab131366) at a dilution of 1/1000.

Lane 1 : NIH/3T3 whole cell lysate 10ug
Lane 2 : ab32058 IP in NIH/3T3 whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab32058 in NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate

All lanes:

Immunoprecipitation - Anti-Sumo 1 antibody [Y299] - ChIP Grade (ab32058)

Predicted band size: 11 kDa

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• WB

Lab

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : Empty
Lane 3 : Sumo 1 knockout HAP1 whole cell lysate (20 μg)
Lane 4 : Empty

Lanes 1 - 4 : Merged signal (red and green). Green - ab32058 observed at 16 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab32058 was shown to react with Sumo 1 in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when Sumo 1 knockout samples were used. Wild-type and Sumo 1 knockout samples were subjected to SDS-PAGE. Samples were incubated with ab32058 and ab8245 (Mouse anti GAPDH loading control) overnight at 4°C at a 1/1000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (ab32058)

Predicted band size: 11 kDa

false

• WB

Lab

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

Blocking and diluting buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (ab32058) at 1/5000 dilution

Lane 1:

HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

A549 (Human lung carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 4:

C6 (Rat glial tumor glial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 11 kDa

false

• WB

Unknown

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (AB32058)

All lanes:

Western blot - Anti-Sumo 1 antibody [Y299] - ChIP Grade (ab32058) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) cell lysate

Lane 2:

NIH/3T3 (Mouse embryo fibroblast cell line) cell lysate

Predicted band size: 11 kDa

Observed band size: 12 kDa,80 kDa

false