RPE65 Antibody (401.8B11.3D9) Summary
Bovine RPE65 microsomal membrane proteins. [UniProt# Q28175]
Cytoplasmic, Microsomeal and Cell Membrane
Retinal Pigment Epithelium Marker
NB100-355 is specific for RPE65.
Protein G purified
Test in a species/application not listed above to receive a full credit towards a future purchase.
- Western Blot 1:500 - 1:1000
- Flow Cytometry
- Immunocytochemistry/Immunofluorescence 1:10-1:500
- Immunohistochemistry 1:250
- Immunohistochemistry-Frozen 1:250
- Immunohistochemistry-Paraffin 1:250-1:500
- Immunocytochemistry 1:250
For Western blot, this antibody has been validated in lysates of bovine RPE membrane and COS7 cells transfected with human RPE65, and in both samples, the antibody recognized a band at ~65 kDa, representing RPE65 protein. Note:- Hamel et al. have reported that in cultured bovine RPE cells, the levels of RPE65 are undetectable in WB after day 14, however, the mRNA levels are detectable by Northern for at least 7 weeks (PMID: 8340400). The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
Based on immunogen sequence similarity, this antibody is predicted to react with Duck/Ostrich and several other Birds species (89%), Zebrafish (74%) and Sea lamprey (71%).
Packaging, Storage & Formulations
Aliquot and store at -20C or -80C. Avoid freeze-thaw cycles.
0.05% Sodium Azide
Protein G purified
Alternate Names for RPE65 Antibody (401.8B11.3D9)
- All-trans-retinyl-palmitate hydrolase
- EC 22.214.171.124
- EC 126.96.36.199
- RBP-binding membrane protein
- Retinal pigment epithelium-specific 65 kDa protein
- retinal pigment epithelium-specific protein (65kD)
- retinal pigment epithelium-specific protein 65kDa
- retinitis pigmentosa 20 (autosomal recessive)
- retinoid isomerohydrolase
- Retinol isomerase
RPE65 (retinal pigment epithelium-specific 65 kDa protein), also called retinoid isomerohydrolase, is an essential retinoid isomerase enzyme of the canonical retinal pigment epithelium/RPE visual cycle and it is highly /preferentially expressed in the RPE layer, monolayer of neuroepithelium-derived cells located between the choroid and photoreceptors of the eye. It is responsible for retinol isomerization (essential reaction required for functional vision) and it converts all-trans retinyl ester to 11-cis retinol. RPE65 protein is functionally conserved throughout the vertebrates, and it is commonly used as an RPE marker by the research community. The loss of RPE65 function has been suggested to involve two distinct mechanisms - loss of catalytic activity and a lower expression level of RPE65. More than 80 different mutations have been reported in this protein and RPE65 gene mutations have been shown to be the cause of Leber's congenital amaurosis (LCA, MIM204000), a severe congenital or early infant-onset form of inherited retinal dystrophy.
This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed
for 1 year from date of receipt.
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Product General Protocols
View specific protocols for RPE65 Antibody (NB100-355):
Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.
FAQs for RPE65 Antibody (NB100-355). (Showing 1 - 5 of 5 FAQs).
I have one question regarding the immunoblot of the RPE65 I always got two bands in positive control. I am not able to understand why. Please clarify my doubt. I am unsing Novus RPE65 antibody.
- RPE65 is subject to several different post-translational modifications that influence its apparent band size in an SDS-PAGE gel. Please see UniProt to learn more. The doublet you are seeing (which we also observe) most likely reflects an unmodified and a modified version of the protein. I hope this helps, but please let me know if you have additional questions.
I am thinking about buying this ab (RPE65 Antibody); you have a nice picture showing how it is working in a saggital retinal section. Could I have the protocol that was used in this labeling? Frozen sections? Antigen retrieval?
- Thank you for your email! We do not have a specific protocol for this Ab. Please see the Immunohistochemistry-Frozen Protocol our lab follows. The recommended dilution for IHC-Fr is 1:250 and antigen retrieval is not neccessary as the the protein is located in the cytoplasm.
We have a question regarding species-specificity for this antibody. Does it work on rat? We'd like to use it on rat eye cryosections. But on the data sheet, only Bovine, Human, Mouse and Porcine were listed.
- We have not tested RPE65 on rat samples. There is 93% similarity between the rat and bovine proteins, which is very promising but I still cannot guarantee that it will work. If you would like to test this antibody in rat samples, you would qualify for the Innovator's Reward Program which rewards our customers for trying our antibodies in previously untested species and applications.
For clarification, is the 1:250 the recommended dilution for only IHC-F or all applications preceding IHC-F on the data sheet? If not, is there a recommended dilution for IHC-P? Do you have any information on antigen retrieval? Citrate buffer or higher pH (EDTA)? Temperature? Time? Regarding incubation time, is 1 hour at room temperature or overnight at 4C recommended?
- The dilution of 1:250 should suffice for IHC-P as well as frozen sections. Unfortunately this item was tested by an outside collaborator who did not provide us with the protocol they used. Generally we recommend using antigen retrieval with citrate buffer and incubating for 1 hour at RT. Some antigens may be tricky and require overnight incubation. As all samples are different, this usually has to be user determined.
Do you recommend an antigen retrieval step for this antibody : RPE65 Antibody (401.8B11.3D9)?. My application is IHC-paraffin cross section!
- Yes, we do recommend antigen retrieval step for NB100-355 and in our IHC validation for this product, heat induced method was employed. You may use citrate buffer (10mM Citric Acid, 0.05% Tween 20, pH 6.0) for this purpose. Please view more information on antigen retrieval protocols.
Positive Control Lysate(s)
Other Available Formats
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Bioinformatics Tool for RPE65 Antibody (NB100-355)
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