Anti-Slit2 antibody [EPR2771]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Slit2 antibody [EPR2771] (AB134166)

Immunohistochemical analysis of paraffin embedded Human fetal kidney tissue labelled with ab134166 at 1/100 dilution.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• WB

Supplier Data

Western blot - Anti-Slit2 antibody [EPR2771] (AB134166)

Blocking buffer and concentration :  5% NFDM/TBST  Diluting buffer and concentration :  5% NFDM/TBST The expression level of Slit2 in T-47D and A549 are consistent with describes in PMID : 24287947 and PMID : 12384551. Negative control : T-47D and A549 (PMID : 24287947 and PMID : 12384551). Slit2 is a secretory protein, so Brefeldin A (BFA) would help to increase the detection of Slit2 in cell lysate. ab181602 was used as a loading control.

All lanes:

Western blot - Anti-Slit2 antibody [EPR2771] (ab134166) at 1/1000 dilution

Lane 1:

HEK-293 (Human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 2:

T-47(Human ductal breast epithelial tumor epithelial cell) whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

Untreated PC-3 (Human prostate adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 5:

PC-3 (Human prostate adenocarcinoma epithelial cell) treated with 300ng/ml Brefeldin A (BFA) for 24 hours 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: 169 kDa

Observed band size: 200 kDa

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Exposure time: 40s

• WB

Lab

Western blot - Anti-Slit2 antibody [EPR2771] (AB134166)

ab134166 was shown to react with SLIT2 in wild-type HAP1 cells in Western blot with loss of signal observed in a SLIT2 knockout cell line. Wild-type HAP1 and SLIT2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 5% milk in TBST for 1 hr before incubation with ab134166 overnight at 4 °C at a 1/1000 dilution. Blots were incubated with secondary antibodies at 0.2 µg/mL before imaging.

These data were provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

All lanes:

Western blot - Anti-Slit2 antibody [EPR2771] (ab134166) at 1/1000 dilution

Lane 1:

Wild-type HAP1 lysate at 40 µg

Lane 2:

SLIT2 knock-out HAP1 lysate at 40 µg

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• WB

Lab

Western blot - Anti-Slit2 antibody [EPR2771] (AB134166)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

ab213204 was used as a His tag loading control.

All lanes:

Western blot - Anti-Slit2 antibody [EPR2771] (ab134166) at 1/1000 dilution

Lane 1:

293T cells transfected with an empty vector containing a His tag whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human Slit1 expression vector containing a His-tag whole cell lysate at 20 µg

Lane 3:

293T cells transfected with a human Slit2 expression vector containing a His-tag whole cell lysate at 20 µg

Lane 4:

293T cells transfected with a human Slit3 expression vector containing a His-tag 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

Observed band size: 250 kDa

false

Exposure time: 1s

• WB

CiteAb

Western blot - Anti-Slit2 antibody [EPR2771] (AB134166)

Slit2 western blot using anti-Slit2 antibody [EPR2771] ab134166. Publication image and figure legend from Zhao, S. J., Shen, Y. F., et al., 2018, Cell Death Dis, PubMed 29523788.

ab134166 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 ab134166 please see the product overview.

c-MYC and PFKFB2 are essential for SLIT2/ROBO1 axis-mediated Warburg effect.a The mRNA expression levels of PFKFBs (PFKFB1, PFKFB2, PFKFB3, and PFKFB4) in sh-Control and sh-ROBO1 U-2OS cells, Values are means ± SD, *p < 0.05, **p < 0.01. b The expression of PFKFBs (PFKFB1, PFKFB2, PFKFB3, and PFKFB4) was detected via real-time polymerase chain reaction (RT-qPCR) in sh-Control and sh-ROBO1 Saos-2 cells. β-actin was used as an internal control in this study. Values are means ± SD, *p < 0.05. c Altered protein expression level of SRC, p-SRC, ERK, p-ERK, c-MYC, and PFKFB2 was detected using western blot upon knockdown of ROBO1 or SLIT2 in OS cells (U-2OS or Saos-2). d A ChIP assay was performed to confirm the potential c-MYC-binding site in the PFKFB2 promoter region in U-2OS and MNNG-HOS cell lines. IgG and input fractions were used as controls. e Luciferase activities of OS cells (U-2OS and Saos-2) in luciferase reporter plasmid containing wild-type and mutant PFKFB2 promoter (mutation site : red). The data shown are mean ± SD, **p < 0.01, ***p < 0.001. f Schematic of SLIT2/ROBO1 axis upregulated PFKFB2 expression through the SRC/ERK/c-MYC pathway in OS cells.

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