KDEL Recombinant Rabbit Monoclonal Antibody [JB42-04]
Catalog# ET7107-86
KDEL Recombinant Rabbit Monoclonal Antibody [JB42-04]
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WB
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IF-Cell
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IF-Tissue
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IHC-P
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FC
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Human
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Mouse
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Rat
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unconjugated
Overview
Product Name
KDEL Recombinant Rabbit Monoclonal Antibody [JB42-04]
Antibody Type
Recombinant Rabbit monoclonal Antibody
Immunogen
Synthetic peptide CSEKDEL.
Species Reactivity
Human, Mouse, Rat
Validated Applications
WB, IF-Cell, IF-Tissue, IHC-P, FC
Molecular Weight
Predicted band size: 25 kDa
Positive Control
A549 cell lysate, HepG2 cell lysate, AGS cell lysate, NIH/3T3 cell lysate, Rat placenta tissue lysate, RAW264.7 cell lysate, C6 cell lysate, Mouse placenta (E16.5) tissue lysate, Mouse lung tissue lysate, Mouse colon tissue lysate, Rat lung tissue lysate, Rat colon tissue lysate, A549, NIH/3T3, C6, human placenta tissue, human small intestine tissue, rat epididymis tissue, human stomach carcinoma tissue, mouse small intestine tissue.
Conjugation
unconjugated
Clone Number
JB42-04
RRID
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
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WB
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1:500-1:2,000
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IF-Cell
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1:100-1:250
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IF-Tissue
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1:400-1:800
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IHC-P
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1:100-1:400
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FC
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1:1,000
Target
Function
Soluble proteins in the endoplasmic reticulum (ER) contain a specific carboxy terminal sequence KDEL (Lys-Asp-Glu-Leu), and include the coat proteins required for vesicle budding from the ER, proteins that form retrograde vesicles on post-ER compartments, and integral membrane proteins that target vesicles to their correct destination. The retention of these soluble proteins in the ER depends on the interaction of the KDEL sequence with the corresponding KDEL receptor, also designated ERD2, in the Golgi apparatus. When KDEL proteins reach the Golgi complex, they are recognized by the KDEL receptor and transported retrograde in COPI-coated vesicles back to the ER. The small GTPase ADP-ribosylation factor 1 (ARF1), a regulator of vesicle transport, interacts with the KDEL receptor. Subsequently, this interaction allows the KDEL receptor to recruit a GTPase-activating protein (GAP) from the cytosol to membranes, which inactivates ARF1.
Background References
1. Majoul I et al. KDEL-cargo regulates interactions between proteins involved in COPI vesicle traffic: measurements in living cells using FRET. Dev Cell 1:139-153 (2001).
2. Cabrera M et al. The retrieval function of the KDEL receptor requires PKA phosphorylation of its C-terminus. Mol Biol Cell 14:4114-4125 (2003).
Sequence Similarity
Belongs to the ERD2 family.
Post-translational Modification
Phosphorylation by PKA at Ser-209 is required for endoplasmic reticulum retention function.
Subcellular Location
Endoplasmic reticulum Membrane.
Synonyms
KDEL antibody
Lys Asp Glu Leu antibody
Images
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Western blot analysis of KDEL on different lysates with Rabbit anti-KDEL antibody (ET7107-86) at 1/1,000 dilution.
Lane 1: A549 cell lysate (20 µg/Lane)
Lane 2: HepG2 cell lysate (20 µg/Lane)
Lane 3: AGS cell lysate (20 µg/Lane)
Lane 4: NIH/3T3 cell lysate (20 µg/Lane)
Lane 5: Rat placenta tissue lysate (20 µg/Lane)
Predicted band size: 25 kDa
Observed band size: 50-100 kDa
Exposure time: Lane 1-4: 1 minute; Lane 5: 3 minutes; 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 (ET7107-86) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. 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 KDEL on different lysates with Rabbit anti-KDEL antibody (ET7107-86) at 1/1,000 dilution.
Lane 1: RAW264.7 cell lysate (20 µg/Lane)
Lane 2: C6 cell lysate (20 µg/Lane)
Lane 3: Mouse placenta (E16.5) tissue lysate (20 µg/Lane)
Predicted band size: 25 kDa
Observed band size: 50-100 kDa
Exposure time: 25 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 (ET7107-86) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. 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 KDEL on different lysates with Rabbit anti-KDEL antibody (ET7107-86) at 1/1,000 dilution.
Lane 1: Mouse lung tissue lysate (20 µg/Lane)
Lane 2: Mouse colon tissue lysate (20 µg/Lane)
Lane 3: Rat lung tissue lysate (20 µg/Lane)
Lane 4: Rat colon tissue lysate (20 µg/Lane)
Predicted band size: 25 kDa
Observed band size: 50-100 kDa
Exposure time: 1 minute; 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 (ET7107-86) at 1/1,000 dilution was used in 5% NFDM/TBST 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 A549 cells labeling KDEL with Rabbit anti-KDEL antibody (ET7107-86) at 1/250 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-KDEL antibody (ET7107-86) at 1/250 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
Immunocytochemistry analysis of NIH/3T3 cells labeling KDEL with Rabbit anti-KDEL antibody (ET7107-86) at 1/250 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-KDEL antibody (ET7107-86) at 1/250 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
Immunocytochemistry analysis of C6 cells labeling KDEL with Rabbit anti-KDEL antibody (ET7107-86) at 1/100 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-KDEL antibody (ET7107-86) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
Immunohistochemical analysis of paraffin-embedded human placenta tissue using anti-KDEL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-86, 1/100) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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Immunohistochemical analysis of paraffin-embedded human small intestine tissue using anti-KDEL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 1% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-86, 1/400) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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Immunohistochemical analysis of paraffin-embedded rat epididymis tissue using anti-KDEL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-86, 1/400) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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Immunohistochemical analysis of paraffin-embedded human stomach carcinoma tissue using anti-KDEL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-86, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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Immunohistochemical analysis of paraffin-embedded mouse small intestine tissue using anti-KDEL antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7107-86, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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Flow cytometric analysis of A549 cells labeling KDEL.
Cells were fixed and permeabilized. Then stained with the primary antibody (ET7107-86, 1/1,000) (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). -
Flow cytometric analysis of NIH/3T3 cells labeling KDEL.
Cells were fixed and permeabilized. Then stained with the primary antibody (ET7107-86, 1/1,000) (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). -
Flow cytometric analysis of C6 cells labeling KDEL.
Cells were fixed and permeabilized. Then stained with the primary antibody (ET7107-86, 1/1,000) (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).
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