Anti-Rab5 antibody - Early Endosome Marker (ab18211)

Overview

  • Product nameAnti-Rab5 antibody - Early Endosome MarkerSee all Rab5 primary antibodies ...
  • Description
    Rabbit polyclonal to Rab5 - Early Endosome Marker
  • Tested applicationsIP, IHC-FrFl, IHC-Fr, ICC/IF, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Dog, Human, Chinese Hamster
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 150 to the C-terminus of Human Rab5.

  • Positive control
    • This antibody gave a positive control in the following human lysate: Jurkat (Human T cell lymphoblast-like cell line) whole cell, NIH 3T3 whole cell, MEF1 whole cell, Brain tissue, Testis tissue, Spinal Cord tissue, PC12 whole cell, Brain tissue and Heart tissue

Properties

Applications

Our Abpromise guarantee covers the use of ab18211 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IP Use at an assay dependent concentration.
IHC-FrFl Use at an assay dependent concentration.
IHC-Fr 1/300.
ICC/IF Use a concentration of 1 µg/ml.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 24 kDa (predicted molecular weight: 24 kDa).Can be blocked with Human Rab5 peptide (ab18625).

Target

Anti-Rab5 antibody - Early Endosome Marker images

  • Anti-Rab5 antibody - Early Endosome Marker (ab18211) at 1 µg/ml + Jurkat cell lysate at 20 µg

    Secondary
    Alexa Fluor Goat polyclonal to Rabbit IgG (700) at 1/5000 dilution

    Predicted band size : 24 kDa
    Observed band size : 24 kDa
    Additional bands at : 40 kDa,70 kDa. We are unsure as to the identity of these extra bands.
  • ICC/IF image of ab18211 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab18211, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

  • All lanes : Anti-Rab5 antibody - Early Endosome Marker (ab18211) at 1 µg/ml

    Lane 1 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab7179)
    Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate (ab46770)
    Lane 3 : Brain (Mouse) Tissue Lysate (ab27253)
    Lane 4 : Testis (Mouse) Tissue Lysate - normal tissue
    Lane 5 : Spinal Cord (Mouse) Tissue Lysate
    Lane 6 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate
    Lane 7 : Brain (Rat) Tissue Lysate - normal tissue
    Lane 8 : Heart (Rat) Tissue Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 24 kDa
    Observed band size : 25 kDa (why is the actual band size different from the predicted?)
  • ab18211 at 1/300 staining adult rat brain (perfusion fixed) tissue sections by IHC-Fr. Adult rat was perfused intracardially with paraformaldehyde 4% in PB 0.2M. The brain was post-fixed in the same fixative for 24 hours. Sections were cryoprotected with sucrose 20% and later frozen in OCT. Sections were incubated in free floating for 12h with the primary antibody (1/300) and later revealed with secondary antibody conjugated with Alexa Fluor ® 488 (1/2000).

    The staining obtained is restricted to the cytoplasm and consists of a small and thin punctate staining. The picture shows the staining obtained at the level of the spinal cord using the X20 objective and zooming on two particular neurons.

    See Abreview

  • ab18211 immunoprecipitated Rab5 in Mouse neuroblastoma N2a whole cell lysate. 10µg of cell lysate was incubaed with primary antibody (1/2000 in dilution buffer:0.025 M Tris, 0.15 M NaCl, 0.001 M EDTA, 1% NP-40, 5% glycerol pH 7.4) for 2 hours at 22°C and an AminoLink® Plus Coupling Resin matrix.
    For Western blotting an HRP conjugated goat monoclonal to rabbit IgG (1/2000) was used.

    See Abreview

  • ICC/IF image of ab18211 stained Hela cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab18211, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • Anti-Rab5 antibody - Early Endosome Marker (ab18211) at 1 µg/ml + Human Rab5 full length protein (ab62956) at 0.001 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Exposure time : 150 seconds

References for Anti-Rab5 antibody - Early Endosome Marker (ab18211)

This product has been referenced in:
  • Suo D  et al. Coronin-1 is a neurotrophin endosomal effector that is required for developmental competition for survival. Nat Neurosci 17:36-45 (2014). WB, ICC/IF ; Rat . Read more (PubMed: 24270184) »
  • Simon NC & Barbieri JT Exoenzyme S ADP-ribosylates Rab5 effector sites to uncouple intracellular trafficking. Infect Immun 82:21-8 (2014). Read more (PubMed: 24101692) »

See all 16 Publications for this product

Product Wall

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing
Sample Mouse Cell lysate - whole cell (hepatocytes)
Specification hepatocytes
Treatment 20 ug/ml poly(I:C)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

Submitted Jul 03 2014

Application Immunocytochemistry/ Immunofluorescence
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: RT°C
Sample Human Cell (Hepatoma)
Specification Hepatoma
Permeabilization Yes - 0.2% Triton X-100
Fixative Formaldehyde
Username

Ms. Alina Deliu

Verified customer

Submitted May 21 2013

Application Immunocytochemistry/ Immunofluorescence
Sample Rat Cell (Rat astrocytes)
Specification Rat astrocytes
Fixative Paraformaldehyde
Permeabilization Yes - 0.3% TritonX in 0.1% PBS
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 24°C
Username

Dr. Ruma Raha-Chowdhury

Verified customer

Submitted Dec 04 2012

Thank you for confirming this information and for your help and cooperation with this case.

As requested, I have asked our accounting department to issue you with a credit note. This can then be redeemed against the invoice of a future order...

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Thank you for your message and for providing this further information.

I am sorry to hear the suggestions made have not improved the results on this occasion. I appreciate the time you have spent on these experiments and would be pleased to ...

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I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement for 2 vials of ab18211.

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Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this...

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Thank you for your reply and for kindly providing this further information and order number.

I look forward to hearing from you with the results from the next experiment.

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Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from three of ourantibodies.

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Application Western blot
Sample Human Cell lysate - whole cell (HepaRG)
Loading amount 50 µg
Specification HepaRG
Gel Running Conditions Reduced Denaturing (12)
Blocking step Milk as blocking agent for 12 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C
Username

Mrs. Alina Macovei

Verified customer

Submitted May 09 2011

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"