Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric)

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized SH-SY5Y (human neuroblastoma epithelial cell) cells labeling RILPL1 with ab307294 at 1/500 dilution, followed by ab98712 Goat Anti-Mouse IgG Fc (DyLight® 488) preadsorbed antibody at 1/1000 dilution (2 µg/ml) (Green). Confocal image showing centrosome and cytoplasmic staining in SH-SY5Y cell line. Low expression : U-937. ab179513 Anti-beta Tubulin rabbit monoclonal antibody was used to counterstain tubulin at 1/200 dilution (10µg/ml) followed by ab150080 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) at a 1/400 dilution (4 µg/ml) (Red). The nuclear counterstain was DAPI (Blue). -ve control 1 : ab307294 at a 1/500 dilution followed by ab150080 at a 1/500 dilution. -ve control 2 : ab179513 at a 1/200 dilution followed by ab98712 at a 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

RILPL1 was immunoprecipitated from 0.35 mg A549 (human lung carcinoma epithelial cell) whole cell lysate 10 µg with ab307294 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307294 at dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : A549 whole cell lysate 10 µg

Lane 2 : ab307294 at 1/30 IP in A549 whole cell lysate 10 µg

Lane 3 : Mouse IgG2a Isotype control (ab18413) instead of ab307294 in A549 whole cell lysate

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

A549 (human lung carcinoma epithelial cell) at 10 µg

Lane 2:

ab307294 at 1/30 IP in A549 (human lung carcinoma epithelial cell) at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 47 kDa

true

Exposure time: 3min

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) NIH/3T3 treated with CFI-400945 ( 500nM ) for18 hours cells labelling PLK4 with ab325938 at 1/50 (10.48 µg/ml) dilution , followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green).

Confocal image showing centrosome staining in NIH/3T3 cells treated with CFI-400945 ( 500nM ) for 18 hours (shown in green), which is co-stained with a centrosome marker. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab307294 Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) was used to counterstain tubulin at 1/100 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized Neuro-2a (mouse neuroblastoma neuroblast) cells labeling RILPL1 with ab307294 at 1/500 dilution, followed by ab98712 Goat Anti-Mouse IgG Fc (DyLight® 488) preadsorbed antibody at 1/1000 dilution (2 µg/ml) (Green). Confocal image showing centrosome staining in Neuro-2a cell line. ab179513 Anti-beta Tubulin rabbit monoclonal antibody was used to counterstain tubulin at 1/200 dilution (10µg/ml) followed by ab150080 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) at a 1/400 dilution (4 µg/ml) (Red). The nuclear counterstain was DAPI (Blue). -ve control 1 : ab307294 at a 1/500 dilution followed by ab150080 at a 1/500 dilution. -ve control 2 : ab179513 at a 1/200 dilution followed by ab98712 at a 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

RILPL1 was immunoprecipitated from 0.35 mg rat brain tissue lysate 10 µg with ab307294 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307294 at dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Rat brain tissue lysate 10 µg

Lane 2 : ab307294 at 1/30 IP in rat brain tissue lysate 10 µg

Lane 3 : Mouse IgG2a Isotype control (ab18413) instead of ab307294 in rat brain tissue lysate

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

Rat brain tissue lysate at 10 µg

Lane 2:

ab307294 at 1/30 IP in rat brain tissue lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 47 kDa

true

Exposure time: 3min

• IP

Supplier Data

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

RILPL1 was immunoprecipitated from 0.35 mg mouse brain tissue lysate 10 µg with ab307294 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307294 at dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Mouse brain tissue lysate 10 µg

Lane 2 : ab307294 at 1/30 IP in mouse brain tissue lysate 10 µg

Lane 3 : Mouse IgG2a Isotype control (ab18413) instead of ab307294 in mouse brain tissue lysate

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Immunoprecipitation - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 10 µg

Lanes 1 - 2:

ab307294 at 1/30 IP in mouse brain tissue lysate at 10 µg

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

Observed band size: 47 kDa

true

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

Blocking and diluting buffer and concentration : 5% NFDM/TBST. In Western blot, ab307294 was shown to bind specifically to RILPL1. A band was observed at 47 kDa in wild-type A549 cell lysates with whereas no signal observed at this size in RILPL1 knockout cell line. Wild-type and RILPL1 knockout A549 cell lysates were kindly provided by Dr. Dario Alessi, University of Dundee. Exposure time : 3 minutes.

All lanes:

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

Wild-type A549 (human lung carcinoma epithelial cell), whole cell lysate 20 μg at 20 µg

Lane 2:

RILPL1 knockout A549 (human lung carcinoma epithelial cell), whole cell lysate 20 μg at 20 µg

Lane 3:

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

Lane 4:

MEF (mouse embryonic fibroblast (immortalized)), whole cell lysate 20 μg at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG Fc (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-fc-hrp-preadsorbed-ab98717'>ab98717</a>) at 1/20000 dilution

Observed band size: 47 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

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

Low expression : U937 (Human Protein Atlas).

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

All lanes:

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

SH-SY5Y (human neuroblastoma epithelial cell), whole cell lysate at 20 µg

Lane 2:

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

Lane 3:

U937 (human histiocytic lymphoma monocyte), whole cell lysate at 20 µg

Lane 4:

Human cerebellum tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG Fc (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-fc-hrp-preadsorbed-ab98717'>ab98717</a>) at 1/20000 dilution

Observed band size: 47 kDa

true

Exposure time: 81s

• WB

Supplier Data

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (AB307294)

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

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

Exposure time :

Lane 1-4 : 180 seconds

Lanes 5-7 : 125 seconds

All lanes:

Western blot - Anti-RILPL1 antibody [MJF-R41-21] - Mouse IgG2a (Chimeric) (ab307294) at 1/1000 dilution

Lane 1:

C6 (rat glial tumor cell), whole cell lysate at 10 µg

Lane 2:

PC-12 (rat adrenal gland pheochromocytoma), whole cell lysate at 10 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast), whole cell lysate at 10 µg

Lane 4:

Rat brain tissue lysate at 10 µg

Lane 5:

Mouse brain tissue lysate at 10 µg

Lane 6:

Mouse heart tissue lysate at 10 µg

Lane 7:

Mouse spleen tissue lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Mouse IgG Fc (HRP) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-fc-hrp-preadsorbed-ab98717'>ab98717</a>) at 1/20000 dilution

Observed band size: 47 kDa

true