Anti-PROX1 antibody [EPR19273]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-PROX1 antibody [EPR19273] (AB199359)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized SH-SY5Y (Human neuroblastoma cell line from bone marrow) and HEK-293 (Human epithelial cell line from embryonic kidney) cells labeling PROX1 with ab199359 at 1/500 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on SH-SY5Y cell line.

Negative expression in HEK-293 cells. HEK-293 doesn't express PROX1. PMID : 18948102.

The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG (AlexaFluor®594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab199359 at 1/500 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2 : ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PROX1 antibody [EPR19273] (AB199359)

Immunohistochemical analysis of paraffin-embedded mouse hippocampus tissue labeling PROX1 with ab199359 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on neurons of mouse hippocampus is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PROX1 antibody [EPR19273] (AB199359)

Immunohistochemical analysis of paraffin-embedded rat hippocampus tissue labeling PROX1 with ab199359 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on neurons of rat hippocampus is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PROX1 antibody [EPR19273] (AB199359)

Immunohistochemical analysis of formalin fixed paraffin embedded mouse brain labelling PROX1 with ab199359 at a concentration of 0.5µg/ml. The immunostaining was performed on a Ventana DISCOVERY ULTRA (Roche Tissue Diagnostics) instrument with a ChromoMap DAB kit and anti-rabbit HQ and anti-HQ HRP detection. Heat mediated antigen retrieval was performed with DISCOVERY cell conditioning solution (CC1) 100°C, pH8.5 for 32mins. ab199359 Anti-PROX1 antibody [EPR19273] was incubated for 16mins at 37°C. Sections were counterstained with Hematoxylin II. Image inset shows absence of staining in secondary antibody only control.

Customers are encouraged to optimise antigen retrieval conditions, antibody concentration, incubation times and temperature for best results in their own IHC assay workflow (automated and manual).

• WB

Supplier Data

Western blot - Anti-PROX1 antibody [EPR19273] (AB199359)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-PROX1 antibody [EPR19273] (ab199359) at 1/1000 dilution

Lane 1:

Human fetal liver lysate at 10 µg

Lane 2:

Human brain lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-PROX1 antibody [EPR19273] (AB199359)

Blocking/Dilution buffer : 5% NFDM/TBST.

HEK-293 doesn't express PROX1. PMID : 18948102.

All lanes:

Western blot - Anti-PROX1 antibody [EPR19273] (ab199359) at 1/1000 dilution

Lane 1:

SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell lysate at 20 µg

Lane 2:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Lane 3:

HEK-293 (Human epithelial cell line from embryonic kidney) 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 1/100000 dilution

Predicted band size: 83 kDa

Observed band size: 83 kDa

false

Exposure time: 10s

• ChIC/CUT&RUN-seq

Supplier Data

ChIC/CUT&RUN sequencing - Anti-PROX1 antibody [EPR19273] (AB199359)

ChIC/CUT&RUN was performed using a pAG-MNAse at a final concentration of 700 ng/mL, 2.5 x 10^5 HepG2 (Human liver hepatocellular carcinoma cell line) cells and 5 µg of ab199359 [EPR19273]. The resulting DNA was sequenced on the Illumina NovaSeq 6000 to a depth of 10 million reads. The negative IgG control ab172730 is also shown.

Additional screenshots of mapped reads can be found in the Protocol booklet in the Product Protocol section.

The University of Geneva owns patents relevant to ChIC (Chromatin Immuno-Cleavage) methods.

• WB

Supplier Data

Western blot - Anti-PROX1 antibody [EPR19273] (AB199359)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1,2 and 3 : minutes; Lane 4 : 5 seconds.

All lanes:

Western blot - Anti-PROX1 antibody [EPR19273] (ab199359) at 1/1000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Rat brain lysate at 10 µg

Lane 3:

Mouse hippocampus lysate at 10 µg

Lane 4:

Rat hippocampus lysate at 10 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 1/100000 dilution

Predicted band size: 83 kDa

Observed band size: 83 kDa

false