Recombinant
RabMAb

Recombinant Anti-Apolipoprotein A I antibody [EP1368Y] (ab52945)

Overview

  • Product name

    Anti-Apolipoprotein A I antibody [EP1368Y]
    See all Apolipoprotein A I primary antibodies
  • Description

    Rabbit monoclonal [EP1368Y] to Apolipoprotein A I
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Sandwich ELISA, IHC-FoFr, WB, IP, Flow Cyt, IHC-P, ICC/IFmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human Apolipoprotein A I aa 1-100 (N terminal). The exact sequence is proprietary.

  • Positive control

    • WB: Human liver and fetal lysates IHC-P: Human liver tissue ICC/IF: HepG2 cells FC: HepG2 cells IP: HepG2 cell lysates, Human fetal liver tissue lysate
  • General notes

    Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.

     

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Applications

Our Abpromise guarantee covers the use of ab52945 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
Sandwich ELISA Use a concentration of 0 - 0.1 mg/ml. Can be paired for Sandwich ELISA with Goat polyclonal to Apolipoprotein A I (HRP) (ab20784).
IHC-FoFr Use at an assay dependent concentration.
WB 1/1000. Detects a band of approximately 31 kDa (predicted molecular weight: 31 kDa).

For unpurified use at 1/20000 dilution.

IP 1/60.
Flow Cyt 1/20.
IHC-P 1/100. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

ICC/IF 1/100 - 1/250.

Target

  • Function

    Participates in the reverse transport of cholesterol from tissues to the liver for excretion by promoting cholesterol efflux from tissues and by acting as a cofactor for the lecithin cholesterol acyltransferase (LCAT). As part of the SPAP complex, activates spermatozoa motility.
  • Tissue specificity

    Major protein of plasma HDL, also found in chylomicrons. Synthesized in the liver and small intestine.
  • Involvement in disease

    Defects in APOA1 are a cause of high density lipoprotein deficiency type 2 (HDLD2) [MIM:604091]; also known as familial hypoalphalipoproteinemia (FHA). Inheritance is autosomal dominant.
    Defects in APOA1 are a cause of the low HDL levels observed in high density lipoprotein deficiency type 1 (HDLD1) [MIM:205400]; also known as analphalipoproteinemia or Tangier disease (TGD). HDLD1 is a recessive disorder characterized by the absence of plasma HDL, accumulation of cholesteryl esters, premature coronary artery disease, hepatosplenomegaly, recurrent peripheral neuropathy and progressive muscle wasting and weakness. In HDLD1 patients, ApoA-I fails to associate with HDL probably because of the faulty conversion of pro-ApoA-I molecules into mature chains, either due to a defect in the converting enzyme activity or a specific structural defect in Tangier ApoA-I.
    Defects in APOA1 are the cause of amyloid polyneuropathy-nephropathy Iowa type (AMYLIOWA) [MIM:107680]; also known as amyloidosis van Allen type or familial amyloid polyneuropathy type III. AMYLIOWA is a hereditary generalized amyloidosis due to deposition of amyloid mainly constituted by apolipoprotein A1. The clinical picture is dominated by neuropathy in the early stages of the disease and nephropathy late in the course. Death is due in most cases to renal amyloidosis. Severe peptic ulcer disease can occurr in some and hearing loss is frequent. Cataracts is present in several, but vitreous opacities are not observed.
    Defects in APOA1 are a cause of amyloidosis type 8 (AMYL8) [MIM:105200]; also known as systemic non-neuropathic amyloidosis or Ostertag-type amyloidosis. AMYL8 is a hereditary generalized amyloidosis due to deposition of apolipoprotein A1, fibrinogen and lysozyme amyloids. Viscera are particularly affected. There is no involvement of the nervous system. Clinical features include renal amyloidosis resulting in nephrotic syndrome, arterial hypertension, hepatosplenomegaly, cholestasis, petechial skin rash.
  • Sequence similarities

    Belongs to the apolipoprotein A1/A4/E family.
  • Post-translational
    modifications

    Palmitoylated.
    Phosphorylation sites are present in the extracelllular medium.
  • Cellular localization

    Secreted.
  • Information by UniProt
  • Database links

  • Alternative names

    • Apo-AI antibody
    • ApoA I antibody
    • ApoA-I antibody
    • APOA1 antibody
    • APOA1_HUMAN antibody
    • Apolipoprotein A-I(1-242) antibody
    • Apolipoprotein A1 antibody
    • Apolipoprotein AI antibody
    • Apolipoprotein of high density lipoprotein antibody
    • ApolipoproteinAI antibody
    • Brp14 antibody
    • high density lipoprotein uptake antibody
    • Ltw1 antibody
    • Lvtw1 antibody
    • MGC117399 antibody
    • Sep1 antibody
    • Sep2 antibody
    see all

Images

  • Anti-Apolipoprotein A I antibody [EP1368Y] (ab52945) at 1/1000 dilution (Purified) + HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

    Predicted band size: 31 kDa
    Observed band size: 31 kDa

  • Anti-Apolipoprotein A I antibody [EP1368Y] (ab52945) at 1/1000 dilution (Purified) + Human liver lysates at 15 µg

    Secondary
    Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

    Predicted band size: 31 kDa
    Observed band size: 31 kDa

  • Anti-Apolipoprotein A I antibody [EP1368Y] (ab52945) at 1/20000 dilution ((unpurified)) + fetal liver lysate at 10 µg

    Secondary
    goat anti-rabbit HRP at 1/2000 dilution

    Predicted band size: 31 kDa
    Observed band size: 31 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human liver tissue sections labeling Apolipoprotein A I with purified ab52945 at 1/100 dilution (1.95 µg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
  • Immunocytochemistry/ Immunofluorescence analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling Apolipoprotein A I with purified ab52945 at 1/250 dilution (0.8 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200 (2.5 µg/ml) dilution. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

  • Flow Cytometry analysis of HepG2 (Human hepatocellular carcinoma epithelial cell) cells labeling Apolipoprotein A I with purified ab52945 at 1/20 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

  • ab52945 (purified) at 1/20 dilution (1ug) immunoprecipitating Apolipoprotein A I in Human fetal liver lysates.
    Lane 1: Human fetal liver lysates 10ug
    Lane 2 (+): ab52945 & Human fetal liver lysates
    Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab52945 in Human fetal liver lysates
    For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.
    Blocking and diluting buffer: 5% NFDM/TBST.

  • Direct ELISA antigen dose-response curve using purified ab52945 at 0-1000ng/mL. Antigen (apo AI peptide) concentration of 1000ng/mL. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) (1/2500) was used as the secondary antibody.

  • ab52945 (unpurified) at 1/100 dilution staining Apolipoprotein A I in human liver by Immunohistochemistry, Paraffin embedded tissue.

  • ab52945 (unpurified) at 1/100 dilution staining Apolipoprotein A I in HEPG2 cells by Immunofluorescence.

References

This product has been referenced in:

  • Camacho J  et al. Brain ApoA-I, ApoJ and ApoE Immunodetection in Cerebral Amyloid Angiopathy. Front Neurol 10:187 (2019). Read more (PubMed: 30918495) »
  • Mazzeo A  et al. Functional analysis of miR-21-3p, miR-30b-5p and miR-150-5p shuttled by extracellular vesicles from diabetic subjects reveals their association with diabetic retinopathy. Exp Eye Res 184:56-63 (2019). Read more (PubMed: 31002820) »
See all 14 Publications for this product

Customer reviews and Q&As

1-7 of 7 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Small Intestine)
Antigen retrieval step
Heat mediated
Permeabilization
No
Specification
Small Intestine
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C
Fixative
Formaldehyde

Arnab Chattopadhyay

Verified customer

Submitted Oct 31 2018

Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Sample
Mouse Tissue sections (Brain)
Permeabilization
Yes - 0.1 M PBS with 1% Triton X
Specification
Brain
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 24°C
Fixative
Paraformaldehyde

Dr. Ruma Raha-Chowdhury

Verified customer

Submitted Feb 05 2016

Application
Western blot
Sample
Mouse Tissue lysate - whole (Brain lysate)
Gel Running Conditions
Reduced Denaturing
Loading amount
30 µg
Specification
Brain lysate
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C

Dr. Ruma Raha-Chowdhury

Verified customer

Submitted Jan 28 2016

Answer

Thank you for your enquiry and interest in our products.

Currently, the blocking peptide for this particular antibody is not available commercially. However, if you wish to check the specificity I would advise to run a no-primary (only secondary antibody) control, a positive control (fetal liver lysate, human liver tissue and HEPG2 cells) and a negative control alongside with the samples.

You may find some useful information on how to find positive control at this site:

https://www.abcam.com/index.html?pageconfig=technicalfaqs#6

If you need any further assistance in the future, please do not hesitate to contact me.

Read More

Answer

Thank you for bringing this to our attention. I agree that your protocol appears correct and that the antibody may be at fault. We sometimes see an improvement in signal for some antibodies after switching the blocking buffer from 5% milk to BSA, but given that you see no signal at all with a dilution of 1/1000, trying a different antibody may be the better approach.

The concentrations of most of our rabbit monoclonal antibodies are not listed on the datasheets because they are usually so low that customers become wary of buying them. These antibodies are unique in having very high affinity for their targets compared to mouse monoclonal antibodies and rabbit polyclonal antibodies. The lot that you have, YI041902C, has an IgG concentration of 0.112 mg/ml.

The only two lots we currently have are the one you have and the one received by the customer you mentioned, who also failed to obtain a western blot signal. If you are not interested in trying a different antibody, I will send a refund. If you would like to try a different antibody, I will be happy to send that instead, such as ab7613, a goat polyclonal which was recently published here: J Biol Chem. 2012 June 1; 287(23): 19341–19345. The western blot is in figure 2, at the following link:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3365970/figure/F2/

Read More

Answer

Thank you for contacting us.

I am sorry that the vial you received was not at the predicted concentration and I apologize for the inconvenience. I have issued a free of charge replacement vial with the order number of xxxxxx.


Please do not hesitate to contact us if you need anything further.

Read More

Answer

Thank you for contacting us. Yes, the Apolipoprotein A I protein ab90760 is the correct target for the antibodies ab52945 and ab20918. However, please note that the only data we have for sELISA are : - ab52945 has been used in sELISA in pair with ab33470, - ab20918 has been used in sELISA in pair with ab64308. See product datasheet for more details : www.abcam.com/ab52945 www.abcam.com/ab33470 www.abcam.com/ab20918 www.abcam.com/ab64308 I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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