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RPE65 Recombinant Rabbit Monoclonal Antibody [JM61-51]

Usd: 385

Specification

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Catalog# ET1703-82

RPE65 Recombinant Rabbit Monoclonal Antibody [JM61-51]

Application

  • WB

  • IP

  • IHC-P

  • IF-Tissue

  • IHC-Fr

Reactivity

  • Human

  • Mouse

  • Rat

Conjugation

  • unconjugated

Overview

Product Name

RPE65 Recombinant Rabbit Monoclonal Antibody [JM61-51]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within Human RPE65 aa 441-484 / 533.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IP, IHC-P, IF-Tissue, IHC-Fr

Molecular Weight

Predicted band size: 61 kDa

Positive Control

Mouse eyeball tissue lysate, Rat eyeball tissue lysate, rat eyeball tissue, mouse eyeball tissue.

Conjugation

unconjugated

Clone Number

JM61-51

RRID

AB_3070437

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

  • 1:2,000

  • IP

  • 1:10-1:50

  • IHC-P

  • 1:1,000

  • IF-Tissue

  • 1:500

  • IHC-Fr

  • 1:500

Target

Function

The retinal pigment epithelium (RPE) is a monolayer simple epithelium in proximity to the outer surface of the retinal photoreceptor cells. Retinal pigment epithelium-specific protein (RPE65) is a 65kDa protein belonging to the beta-carotene dioxygenase family. This protein is important in 11-cis retinal production as well as in visual pigment regeneration. RPE65 is attached to the membrane by a lipid anchor when palmitoylated (membrane form) and soluble when unpalmitoylated. The soluble form of the protein binds vitamin A. Defects in RPE65 causes autosomal dominant retinitis pigmentosa and/or Leber congenital amaurosis type 2.

Background References

1. Zhou J et al. Correlations between Photodegradation of Bisretinoid Constituents of Retina and Dicarbonyl Adduct Deposition. J Biol Chem 290:27215-27 (2015).

2. Sukseree S et al. Tyrosinase-Cre-Mediated Deletion of the Autophagy Gene Atg7 Leads to Accumulation of the RPE65 Variant M450 in the Retinal Pigment Epithelium of C57BL/6 Mice. PLoS One 11:e0161640 (2016).

Sequence Similarity

Belongs to the carotenoid oxygenase family.

Tissue Specificity

Retina (at protein level). Retinal pigment epithelium specific.

Post-translational Modification

Palmitoylation by LRAT regulates ligand binding specificity; the palmitoylated form (membrane form) specifically binds all-trans-retinyl-palmitate, while the soluble unpalmitoylated form binds all-trans-retinol (vitamin A) (By similarity).

Subcellular Location

Cytoplasm. Cell membrane.

UNIPROT

SwissProt: Q16518 Human

SwissProt: Q91ZQ5 Mouse

SwissProt: O70276 Rat

Synonyms

All-trans-retinyl-palmitate hydrolase antibody

LCA 2 antibody

LCA2 antibody

Leber congenital amaurosis antibody

mRPE 65 antibody

mRPE65 antibody

p63 antibody

rd 12 antibody

rd12 antibody

Retinal pigment epithelium specific 61 kDa protein antibody

Expand

Images

  • Western blot analysis of RPE65 on different lysates with Rabbit anti-RPE65 antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/2,000 dilution.<br /><br />Lane 1: Mouse eyeball tissue lysate<br />Lane 2: Rat eyeball tissue lysate<br /><br />Lysates/proteins at 40 µg/Lane.<br /><br />Predicted band size: 61 kDa<br />Observed band size: 61 kDa<br /><br />Exposure time: 59 seconds; ECL: K1801;<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/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/2,000 dilution was used in 5% NFDM/TBST 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.

    Western blot analysis of RPE65 on different lysates with Rabbit anti-RPE65 antibody (ET1703-82) at 1/2,000 dilution.

    Lane 1: Mouse eyeball tissue lysate
    Lane 2: Rat eyeball tissue lysate

    Lysates/proteins at 40 µg/Lane.

    Predicted band size: 61 kDa
    Observed band size: 61 kDa

    Exposure time: 59 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 (ET1703-82) at 1/2,000 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.

  • Immunohistochemical analysis of paraffin-embedded rat eyeball tissue with Rabbit anti-RPE65 antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/1,000 dilution.<br /><br />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 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/1,000 dilution for 1 hour 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.

    Immunohistochemical analysis of paraffin-embedded rat eyeball tissue with Rabbit anti-RPE65 antibody (ET1703-82) at 1/1,000 dilution.

    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 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1703-82) at 1/1,000 dilution for 1 hour 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.

  • Immunohistochemical analysis of paraffin-embedded mouse eyeball tissue with Rabbit anti-RPE65 antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/1,000 dilution.<br /><br />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 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/1,000 dilution for 1 hour 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.

    Immunohistochemical analysis of paraffin-embedded mouse eyeball tissue with Rabbit anti-RPE65 antibody (ET1703-82) at 1/1,000 dilution.

    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 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1703-82) at 1/1,000 dilution for 1 hour 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.

  • Immunofluorescence analysis of paraffin-embedded mouse eyeball tissue labeling RPE65 with Rabbit anti-RPE65 antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>) at 1/500 dilution.<br /><br />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 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (<a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>, green) at 1/500 dilution overnight at 4 ℃, washed with PBS. 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. Nuclei were counterstained with DAPI (blue).

    Immunofluorescence analysis of paraffin-embedded mouse eyeball tissue labeling RPE65 with Rabbit anti-RPE65 antibody (ET1703-82) at 1/500 dilution.

    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 10% negative goat serum for 1 hour at room temperature, washed with PBS, and then probed with the primary antibody (ET1703-82, green) at 1/500 dilution overnight at 4 ℃, washed with PBS. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. Nuclei were counterstained with DAPI (blue).

  • Application: Immunofluorescence (IHC-Fr)<br /><br />Species: Mouse<br />Tissue: Eyeball<br />Sample: Frozen section<br /><br />Antigen retrieval: Not required<br /><br />Wash buffer: 1× TBST<br />Blocking: 10% normal goat serum + 1% Triton X-100 + 0.3 M Glycine in TBST, 30 minutes at room temperature.<br /><br />Primary antibody: <a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>, 1/500, overnight at 4℃.<br />Secondary antibody: Goat Anti-Rabbit IgG (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>), 1.5 hours at room temperature.

    Application: Immunofluorescence (IHC-Fr)

    Species: Mouse
    Tissue: Eyeball
    Sample: Frozen section

    Antigen retrieval: Not required

    Wash buffer: 1× TBST
    Blocking: 10% normal goat serum + 1% Triton X-100 + 0.3 M Glycine in TBST, 30 minutes at room temperature.

    Primary antibody: ET1703-82, 1/500, overnight at 4℃.
    Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 1.5 hours at room temperature.

  • Application: Immunofluorescence (IHC-Fr)<br /><br />Species: Rat<br />Tissue: Eyeball<br />Sample: Frozen section<br /><br />Antigen retrieval: Not required<br /><br />Wash buffer: 1× TBST<br />Blocking: 10% normal goat serum + 1% Triton X-100 + 0.3 M Glycine in TBST, 30 minutes at room temperature.<br /><br />Primary antibody: <a href="/products/ET1703-82" style="font-weight: bold;text-decoration: underline;">ET1703-82</a>, 1/500, overnight at 4℃.<br />Secondary antibody: Goat Anti-Rabbit IgG (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>), 1.5 hours at room temperature.

    Application: Immunofluorescence (IHC-Fr)

    Species: Rat
    Tissue: Eyeball
    Sample: Frozen section

    Antigen retrieval: Not required

    Wash buffer: 1× TBST
    Blocking: 10% normal goat serum + 1% Triton X-100 + 0.3 M Glycine in TBST, 30 minutes at room temperature.

    Primary antibody: ET1703-82, 1/500, overnight at 4℃.
    Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 1.5 hours at room temperature.

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