Anti-PAR2 antibody [EPR13675]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAR2 antibody [EPR13675] (AB180953)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue sections labeling PAR2 with Purified ab180953 at 1/100 dilution (1.38 µg/mL). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-PAR2 antibody [EPR13675] (AB180953)

Flow cytometry overlay histogram showing left HT-29 positive cells and right negative Daudi stained with ab180953 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10μg/ml human IgG and 10% normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody (ab180953) (1x 106 in 100μl at 5.0μg/ml (1/560)) for 30min at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C

Isotype control antibody (black line) was Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-PAR2 antibody [EPR13675] (AB180953)

Intracellular Flow Cytometry analysis of MCF-7 (human breast carcinoma) cells labeling PAR2 with unpurified ab180953 at 1/20 dilution (10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-PAR2 antibody [EPR13675] (AB180953)

Immunocytochemistry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling PAR2 with Purified ab180953 at 1/50 dilution (2.76 µg/mL). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) 1/200 dilution (2.5 µg/mL). Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 dilution (2 µg/mL). DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAR2 antibody [EPR13675] (AB180953)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue sections labeling PAR2 with Purified ab180953 at 1/100 dilution (1.38 µg/mL). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PAR2 antibody [EPR13675] (AB180953)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse kidney tissue sections labeling PAR2 with Purified ab180953 at 1/100 dilution (1.38 µg/mL). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• WB

Unknown

Western blot - Anti-PAR2 antibody [EPR13675] (AB180953)

We are unsure how to define the extra bands.

All lanes:

Western blot - Anti-PAR2 antibody [EPR13675] (ab180953) at 1/1000 dilution

Lane 1:

Human kidney lysate at 15 µg

Lane 2:

Mouse kidney lysate at 15 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 44 kDa

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• WB

Unknown

Western blot - Anti-PAR2 antibody [EPR13675] (AB180953)

We are unsure how to define the extra bands.

All lanes:

Western blot - Anti-PAR2 antibody [EPR13675] (ab180953) at 1/1000 dilution

Lane 1:

HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysate at 15 µg

Lane 2:

Rat kidney lysate at 15 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: 44 kDa

false

• WB

CiteAb

Western blot - Anti-PAR2 antibody [EPR13675] (AB180953)

PAR2 western blot using anti-PAR2 antibody [EPR13675] ab180953. Publication image and figure legend from Klinngam, W., Fu, R., et al., 2018, Int J Mol Sci, PubMed 30423938.

ab180953 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab180953 please see the product overview.

PAR-2 gene and protein expression after 48 h of PAR-2 or scrambled siRNA transfection in human corneal epithelial cells. (A) PAR-2 gene expression in HCE-T cells transfected with PAR-2 or scrambled siRNA. PAR-2 gene expression was normalized to expression of the endogenous gene, GAPDH (n = 3 samples/group; (B) PAR-2 bands measured by Western Blot in lysates from human corneal epithelial cells transfected with PAR-2 or scrambled siRNA; (C) PAR-2 band intensity in human corneal epithelial cells transfected with PAR-2 or scrambled siRNA. The intensity signal of PAR-2 band was normalized to the band intensity of GAPDH and designated as 100% for scrambled siRNA-treated cells (n = 5 samples/group); (D) PAR-2 protein expression in HCE-T cells transfected with PAR-2 or scrambled siRNA as determined by ELISA. PAR-2 protein expression was normalized to total protein in lysates (n = 3 samples in PAR-2 siRNA transfected and n = 2 in scrambled siRNA transfected, * p ≤ 0.05, *** p ≤ 0.001 data are represented as mean ± SEM, and a two-tailed, unpaired Student’s t-test was used to compare PAR-2 siRNA transfected to scrambled siRNA transfected cells).

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