Invitrogen

APOE Recombinant Rabbit Monoclonal Antibody (16H22L18)

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This Antibody was verified by Relative expression to ensure that the antibody binds to the antigen stated. View Details

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APOE Antibody in Immunofluorescence (IF) — Immunofluorescent analysis of ApoE in HepG2 cells using an ApoE recombinant rabbit monoclonal antibody (Product # 701241) followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody (green) (Image A). Nuclei were stained using DAPI (Image B) and actin stained with Alexa Fluor 594 phalloidin (red) (image C). Image D is a composite image showing cytoplasmic localization of ApoE.

APOE Antibody in Immunocytochemistry (ICC) — Immunofluorescence analysis of APOE was performed using 70% confluent log phase Hep G2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with APOE Recombinant Rabbit Monoclonal Antibody (16H22L18) (Product # 701241) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034), (1:2000), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b: Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing Cytoplasmic localization. Panel e represents A-431 showing no expression of APOE. Panel f represents control Hep G2 cells with no primary antibody to assess background. The images were captured at 60X magnification.

APOE Antibody in Western Blot (WB) — Western blot was performed using Anti-APOE Recombinant Rabbit Monoclonal Antibody (16H22L18) (Product # 701241) and a 34 kDa band corresponding to APOE was observed across the cell lines and tissues tested. Whole cell extracts (30 µg lysate) of Hep G2 (Lane 1), Hep G2 treated with Protein Transport Inhibitor (PTI; 1x for 4 hours) (Lane 2), Caco-2 (Lane 3), Caco-2 treated with Protein Transport Inhibitor (PTI; 1x for 4 hours) (Lane 4), A-431 (Lane 5), A-431 treated with Protein Transport Inhibitor (PTI; 1x for 4 hours) (Lane 6), Daudi (Lane 7), Daudi treated with Protein Transport Inhibitor (PTI; 1x for 4 hours) (Lane 8), Mouse Skeletal Muscle (Lane 9), Mouse Liver (Lane 10), Rat Liver (Lane 11) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (2 ug/ml) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036,1:10000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

APOE Antibody in Western Blot (WB) — Western blot analysis was performed on Fig 1: Human plasma (Lane 1); Fig 2: Whole cell extracts (30 µg lysate) of Hep G2 (Lane 1) and Caco-2 (Lane 2). The blots were probed with Anti-ApoE Rabbit Monoclonal Antibody (Product # 701241, 2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). In Fig 1, a 36 kDa band corresponding to ApoE was observed. In Fig 2, a 34 kDa band was observed across both cell lines, and, an additional 36 kDa band corresponding to an isoform of ApoE was observed in Lane 1. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®12 % Bis-Tris gel (Product # NP0342BOX), XCell SureLock Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

APOE Antibody in Western Blot (WB) — Western blot analysis of ApoE in mouse brain (lane 1) and rat liver lysate (lane 2) using an ApoE recombinant rabbit monoclonal antibody (Product # 701241) at a dilution of 2 µg/mL. Samples were detected using chemiluminescence (ECL). Results show a band at ~34kDa.

APOE Antibody in Relative expression — Antibody specificity was demonstrated by detection of differential basal expression of the target across cell models owing to their inherent genetic constitution. Immunofluorescence analysis using Anti-APOE Recombinant Rabbit Monoclonal Antibody (16H22L18) (Product # 701241), shows expression of APOE only in Hep G2 cells as compared to A-431 cells. {RE}

APOE Antibody in Relative expression — Antibody specificity was demonstrated by detection of differential basal expression of the target across cell lines tested owing to their inherent genetic constitution. Relative expression of APOE was observed in Hep G2, CaCo-2 as compared to A-431, Daudi using Anti-APOE Recombinant Rabbit Monoclonal Antibody (16H22L18) (Product # 701241) in Western Blot. {RE}

APOE Antibody — Fig. 4 Natural AbetaOs induced amyloidogenic processing in astrocytes. Western blots and quantitative analysis showing BACE1, sAPPbeta and apoE levels in primary astrocyte (a) and neuron (e) cultures. Bar graphs illustrating the protein expression levels of BACE1, sAPPbeta and apoE in primary astrocyte (b-d) and neuron (f-h) (one-way ANOVA, b: F = 52.03, p < .0001, c: F = 69.79, p < .0001, d: F = 46.81, p < .0001, f: F = 0.436, p < .729, g: F = 0.143, p = .933, h: F = 0.126, p = .943, n = 6/group). The levels of Abeta40 and Abeta42 in the medium obtained from astrocytes or neurons (i-l) (one-way ANOVA, i: F = 310, p < .0001, j: F = 53.06, p < .0001, k: F = 2.73, p = .051, l: F = 2.11, p = .132, n = 6/group). The data are shown as the mean +- SEM; * p < .05, ** p < .01. Fig. 4

APOE Antibody — Fig. 5 Natural AbetaOs injection induced behavioral and biochemical changes in SD rats. (a) MWM escape latency during training (one-way ANOVA, F = 29.34, p < .0001; n = 10/group). (b) Time in the target quadrant (one-way ANOVA, F = 14.79, p < .0001, n = 10/group). (c) The number of crossings (one-way ANOVA, F = 9.81, p = .0006, n = 10/group). (d) Western blot showing dynamic changes in Abeta assembly (4G8) in the brains of SD rats (time indicated above the lanes). The right lanes indicate the respective migration positions of monomers, trimers, nonamers, dodecamers and sAPP. (e-h) Bar graphs illustrating the quantitative analysis of monomers, trimers, nonamers and dodecamers (one-way ANOVA, e: F = 108.3, p < .0001; f: F = 164.9, p < .0001; g: F = 45.89, p < .0001; h: F = 61.43, p < .0001; n = 6/group). (i) Western blots showing the expression of GFAP, BACE1, sAPPbeta and apoE in rat brains. (j-m) Bar graphs illustrating the quantitative analysis of the Western blot results (one-way ANOVA, j: F = 61.83, p < .0001; k: F = 31.09, p < .0001; l: F = 41.37, p < .0001; m: F = 72.38, p < .0001; n = 6/group). The data are shown as the mean +- SEM; * p < .05, ** p < .01. Fig. 5

Product Details

Applications

Tested Dilution

Publications

Immunocytochemistry (ICC)

1-3 µg/mL

-

Immunofluorescence (IF)

1-3 µg/mL

-

Western Blot (WB)

2-4 µg/mL

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Product Specifications

Species Reactivity

Human, Mouse, Rat

Published species

Human

Host / Isotype

Rabbit / IgG

Class

Recombinant Monoclonal

Type

Antibody

Clone

16H22L18

Immunogen

Peptide corresponding to amino acids 240–251 of human ApoE

Conjugate

Unconjugated

Form

Liquid

Concentration

0.5 mg/mL

Purification

Protein A

Storage buffer

PBS

Contains

0.09% sodium azide

Storage conditions

Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.

RRID

AB_2532438

Product Specific Information

This antibody is predicted to react with non-human primate and mouse based on sequence homology.

Intact IgG appears on a non-reducing gel as ~150 kDa band and upon reduction generating a ~25 kDa light chain band and a ~50 kDa heavy chain.

Recombinant rabbit monoclonal antibodies are produced using in vitro expression systems. The expression systems are developed by cloning in the specific antibody DNA sequences from immunoreactive rabbits. Then, individual clones are screened to select the best candidates for production. The advantages of using recombinant rabbit monoclonal antibodies include: better specificity and sensitivity, lot-to-lot consistency, animal origin-free formulations, and broader immunoreactivity to diverse targets due to larger rabbit immune repertoire.

Target Information

Apo E (Apolipoprotein E) plays an important role in the metabolism of lipids in the plasma, and is also is a constituent of various plasma lipoprotein-lipid particles. The protein is synthesized in liver, brain, spleen, and the kidney, and concentrations are high in interstitial fluids, where it plays a role in distributing cholesterol from cells where cholesterol is in excess to cells which require cholesterol. Apo E also plays an important role in the formation of very low density lipoprotein and chylomicrons. The Apo E gene is located on chromosome 19 at position 19q13. 2.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Bioinformatics

Protein Aliases: Apo-E; ApoE4; Apolipoprotein; Apolipoprotein E; apolipoprotein E3; apolipoprotein E4; B2G1; BG; MGC1571

Gene Aliases: AD2; AI255918; APO-E; APOE; APOEA; LDLCQ5; LPG

UniProt ID: (Human) P02649, (Mouse) P08226, (Rat) P02650

Entrez Gene ID: (Human) 348, (Mouse) 11816, (Rat) 25728

response to reactive oxygen species retinoid metabolic process negative regulation of endothelial cell proliferation response to dietary excess triglyceride metabolic process cholesterol catabolic process cellular calcium ion homeostasis receptor-mediated endocytosis cytoskeleton organization G-protein coupled receptor signaling pathway nitric oxide mediated signal transduction synaptic transmission, cholinergic cholesterol metabolic process regulation of gene expression negative regulation of platelet activation positive regulation of cholesterol esterification positive regulation of cholesterol efflux long-chain fatty acid transport protein import virion assembly triglyceride catabolic process cGMP-mediated signaling negative regulation of blood coagulation regulation of axon extension positive regulation of cGMP biosynthetic process neuron projection regeneration regulation of Cdc42 protein signal transduction positive regulation of low-density lipoprotein particle receptor catabolic process cholesterol efflux phospholipid efflux very-low-density lipoprotein particle remodeling low-density lipoprotein particle remodeling high-density lipoprotein particle remodeling high-density lipoprotein particle assembly chylomicron remnant clearance high-density lipoprotein particle clearance very-low-density lipoprotein particle clearance lipoprotein metabolic process lipoprotein biosynthetic process lipoprotein catabolic process vasodilation cholesterol homeostasis negative regulation of MAP kinase activity negative regulation of neuron apoptotic process negative regulation of blood vessel endothelial cell migration reverse cholesterol transport positive regulation by host of viral process negative regulation of cholesterol biosynthetic process positive regulation of lipid biosynthetic process intracellular transport regulation of neuronal synaptic plasticity artery morphogenesis negative regulation of inflammatory response positive regulation of nitric-oxide synthase activity positive regulation of membrane protein ectodomain proteolysis maintenance of location in cell fatty acid homeostasis positive regulation of dendritic spine development negative regulation of canonical Wnt signaling pathway AMPA glutamate receptor clustering NMDA glutamate receptor clustering cellular oxidant detoxification regulation of beta-amyloid clearance negative regulation of neuron death positive regulation of postsynaptic membrane organization negative regulation of presynaptic membrane organization negative regulation of beta-amyloid formation positive regulation of dendritic spine maintenance positive regulation of phospholipid efflux positive regulation of lipid transport across blood brain barrier lipid metabolic process cholesterol biosynthetic process lipid transport response to oxidative stress aging protein localization peripheral nervous system axon regeneration axon regeneration regulation of cholesterol transport response to retinoic acid response to insulin positive regulation of catalytic activity response to ethanol positive regulation of axon extension oligodendrocyte differentiation negative regulation of lipid biosynthetic process lipid homeostasis negative regulation of dendritic spine development cellular response to interleukin-1 cellular response to growth factor stimulus cellular response to cholesterol cardiovascular system development negative regulation of cholesterol efflux regulation of neuron death positive regulation of neuron death negative regulation of postsynaptic membrane organization positive regulation of presynaptic membrane organization positive regulation of beta-amyloid formation regulation of tau-protein kinase activity negative regulation of dendritic spine maintenance positive regulation of neurofibrillary tangle assembly negative regulation of phospholipid efflux negative regulation of lipid transport across blood brain barrier

Process(es)

beta-amyloid binding lipid transporter activity protein binding phospholipid binding heparin binding lipid binding cholesterol binding antioxidant activity cholesterol transporter activity identical protein binding protein homodimerization activity metal chelating activity tau protein binding low-density lipoprotein particle receptor binding phosphatidylcholine-sterol O-acyltransferase activator activity very-low-density lipoprotein particle receptor binding lipoprotein particle binding receptor binding hydroxyapatite binding