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RAB8A Recombinant Rabbit Monoclonal Antibody [JE64-79]

Usd: 385

Specification

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Catalog# HA721127

RAB8A Recombinant Rabbit Monoclonal Antibody [JE64-79]

Application

  • WB

  • FC

  • IF-Cell

  • IP

Reactivity

  • Human

  • Mouse

  • Rat

Conjugation

  • unconjugated

Overview

Product Name

RAB8A Recombinant Rabbit Monoclonal Antibody [JE64-79]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within human RAB8A aa 151-200/207.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, FC, IF-Cell, IP

Molecular Weight

Predicted band size: 24 kDa

Positive Control

Hela cell lysate, HepG2 cell lysate, NIH/3T3 cell lysate, RAW264.7 cell lysate, C2C12 cell lysate, L6 cell lysate, HeLa.

Conjugation

unconjugated

Clone Number

JE64-79

RRID

AB_3072251

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:1,000

  • FC

  • 1:500-1:1,000

  • IF-Cell

  • 1:100

  • IP

  • 1-2μg/sample

Target

Function

The protein encoded by this gene is a member of the RAS superfamily which are small GTP/GDP-binding proteins with an average size of 200 amino acids. The RAS-related proteins of the RAB/YPT family may play a role in the transport of proteins from the endoplasmic reticulum to the Golgi and the plasma membrane. This protein shares 97%, 96%, and 51% similarity with the dog RAB8, mouse MEL, and mouse YPT1 proteins, respectively and contains the 4 GTP/GDP-binding sites that are present in all the RAS proteins. The putative effector-binding site of this protein is similar to that of the RAB/YPT proteins. However, this protein contains a C-terminal CAAX motif that is characteristic of many RAS superfamily members but which is not found in YPT1 and the majority of RAB proteins. Although this gene was isolated as a transforming gene from a melanoma cell line, no linkage between MEL and malignant melanoma has been demonstrable. This oncogene is located 800 kb distal to MY09B on chromosome 19p13.1.

Background References

1. Tong SJ. et. al. Guanine nucleotide exchange factors activate Rab8a for Toll-like receptor signalling. Small GTPases. 2021 Jan

2. Wall AA. et. al. Rab8a localisation and activation by Toll-like receptors on macrophage macropinosomes. Philos Trans R Soc Lond B Biol Sci. 2019 Feb

Subcellular Location

Cell membrane, golgi apparatus, recycling endosome membrane, cytoplasm, centriole, cilium basal body, cilium axoneme, cilium, phagosome, phagosome membrane, midbody.

UNIPROT

SwissProt: P61006 Human

SwissProt: P55258 Mouse

SwissProt: P35280 Rat

Synonyms

AA409338 antibody

MEL antibody

Mel transforming oncogene (derived from cell line NK14) RAB8 homolog antibody

Mel transforming oncogene (derived from cell line NK14) antibody

Mel transforming oncogene (RAB8 homolog) antibody

Mel transforming oncogene antibody

MGC124948 antibody

Oncogene c mel antibody

Oncogene c-mel antibody

RAB8 antibody

Expand

Images

  • Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/1,000 dilution.<br /><br />Lane 1: Hela cell lysate<br />Lane 2: HepG2 cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 24 kDa<br />Observed band size: 24 kDa<br /><br />Exposure time: 2 minutes;<br /><br />15% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/1,000 dilution.

    Lane 1: Hela cell lysate
    Lane 2: HepG2 cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 24 kDa
    Observed band size: 24 kDa

    Exposure time: 2 minutes;

    15% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721127) at 1/1,000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/500 dilution.<br /><br />Lane 1: NIH/3T3 cell lysate<br />Lane 2: RAW264.7 cell lysate<br />Lane 3: C2C12 cell lysate<br />Lane 4: L6 cell lysate<br /><br />Lysates/proteins at 30 µg/Lane.<br /><br />Predicted band size: 24 kDa<br />Observed band size: 24 kDa<br /><br />Exposure time: 2 minutes 16 seconds;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:100,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/500 dilution.

    Lane 1: NIH/3T3 cell lysate
    Lane 2: RAW264.7 cell lysate
    Lane 3: C2C12 cell lysate
    Lane 4: L6 cell lysate

    Lysates/proteins at 30 µg/Lane.

    Predicted band size: 24 kDa
    Observed band size: 24 kDa

    Exposure time: 2 minutes 16 seconds;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721127) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

  • <span style="font-weight: bold;">☑ Knockdown (KD)</span><br /><br />Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/1,000 dilution.<br /><br />Lane 1: HAP1-parental cell lysate<br />Lane 2: HAP1-RAB8A KD cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 24 kDa<br />Observed band size: 24 kDa<br /><br />Exposure time: 30 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/1,000 dilution was used in <a href="/products/K1803" style="font-weight: bold;text-decoration: underline;">K1803</a> at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Knockdown (KD)

    Western blot analysis of RAB8A on different lysates with Rabbit anti-RAB8A antibody (HA721127) at 1/1,000 dilution.

    Lane 1: HAP1-parental cell lysate
    Lane 2: HAP1-RAB8A KD cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 24 kDa
    Observed band size: 24 kDa

    Exposure time: 30 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721127) at 1/1,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunocytochemistry analysis of HeLa cells labeling RAB8A with Rabbit anti-RAB8A antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-RAB8A antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>) at 1/100 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of HeLa cells labeling RAB8A with Rabbit anti-RAB8A antibody (HA721127) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-RAB8A antibody (HA721127) at 1/100 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Flow cytometric analysis of HeLa cells labeling RAB8A.<br /><br />Cells were fixed and permeabilized. Then stained with the primary antibody (<a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a>, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor&trade; 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (<a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

    Flow cytometric analysis of HeLa cells labeling RAB8A.

    Cells were fixed and permeabilized. Then stained with the primary antibody (HA721127, 1ug/ml) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

  • RAB8A was immunoprecipitated from 0.2 mg HeLa cell lysate with <a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a> at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using <a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a> at 1/1,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.<br /><br />Lane 1: HeLa cell lysate (input)<br />Lane 2: <a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a> IP in HeLa cell lysate<br />Lane 3: Rabbit IgG instead of <a href="/products/HA721127" style="font-weight: bold;text-decoration: underline;">HA721127</a> in HeLa cell lysate<br /><br />Blocking/Dilution buffer: 5% NFDM/TBST<br />Exposure time: 4 seconds; ECL: <a href="/products/K1801" style="font-weight: bold;text-decoration: underline;">K1801</a>

    RAB8A was immunoprecipitated from 0.2 mg HeLa cell lysate with HA721127 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA721127 at 1/1,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

    Lane 1: HeLa cell lysate (input)
    Lane 2: HA721127 IP in HeLa cell lysate
    Lane 3: Rabbit IgG instead of HA721127 in HeLa cell lysate

    Blocking/Dilution buffer: 5% NFDM/TBST
    Exposure time: 4 seconds; ECL: K1801

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