Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized A549 (human lung carcinoma cell line) cell line labeling RAB8A with ab241061 at 1/600 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody. This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Immunofluorescent analysis of 3% paraformaldehyde-fixed, 0.1% Saponin permeabilized A549 (human lung carcinoma cell line) and RAB8A knock-out A549 cells labeling RAB8A with ab241061 at 1/100 dilution.

This image is kindly provided by our collaborator Dr. Dario Alessi, University of Dundee.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized A549 (human lung carcinoma cell line) cell line labeling RAB8A with ab241061 at 1/600 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) at 1/2000 dilution was used as the secondary antibody.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human epithelial cell line from cervix adenocarcinoma) cells labeling RAB8A with ab241061 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining in HeLa cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A549 (human lung carcinoma cell line) cells labeling RAB8A with ab241061 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasmic staining in A549 cells.

The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) (red) at 1/200 dilution.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Immunohistochemical analysis of paraffin-embedded human bladder cancer tissue labeling RAB8A with ab241061 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on human bladder cancer (PMID : 14581456). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

Immunohistochemical analysis of paraffin-embedded human breast tissue labeling RAB8A with ab241061 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on human breast (PMID : 14581456). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

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

• IP

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Immunoprecipitation - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

RAB8A was immunoprecipitated from 0.35 mg of A549 (human lung carcinoma cell line) whole cell lysate with ab241061 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab241061 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5000 dilution.

Lane 1 : A549 whole cell lysate 10 μg (Input).

Lane 2 : ab241061 IP in A549 whole cell lysate .

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab241061 in A549 whole cell lysate .

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

Exposure time : 8 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab241061).

All lanes:

Immunoprecipitation - Anti-RAB8A antibody [MJF-R22-79-3] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-mjf-r22-79-3-ab241061'>ab241061</a>)

Predicted band size: 24 kDa

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• WB

Supplier Data

Western blot - Anti-RAB8A antibody [MJF-R22-79-3] - BSA and Azide free (AB243568)

This data was developed using the same antibody clone in a different buffer formulation (ab241061).

Lanes 1-4 : Merged signal (red and green). Green - ab241061 observed at 24 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab241061 Anti-RAB8A antibody [MJF-R22-79-3] was shown to specifically react with RAB8A in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264993 (knockout cell lysate ab257195) was used. Wild-type and RAB8A knockout samples were subjected to SDS-PAGE. ab ab241061 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-RAB8A antibody [MJF-R22-79-3] (<a href='/en-us/products/primary-antibodies/rab8a-antibody-mjf-r22-79-3-ab241061'>ab241061</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

RAB8A knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human RAB8A knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-rab8a-knockout-hela-cell-line-ab264993'>ab264993</a>)

Lane 3:

HCT116 cell lysate at 20 µg

Lane 4:

Human brain tissue lysate at 20 µg

Predicted band size: 24 kDa

Observed band size: 24 kDa

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