Immunofluorescent analysis of Early Endosomal Antigen 1 was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with Early Endosomal Antigen 1 Rabbit Polyclonal Antibody (PA1-063A) at 2ug/ml in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (A27034) a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
|Tested species reactivity||Human, Mouse|
|Published species reactivity||Rat, Human, Mouse|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues T(1391) P S S K K P V R V C D A C F N D L Q G(1410) of human EEA1.|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS with 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Immunocytochemistry (ICC)||2 µg/mL|
|Immunofluorescence (IF)||2 µg/mL|
|Western Blot (WB)||2 µg/mL|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
PA1-063A detects early endosomal antigen 1 (EEA1) from mouse and human samples. Previous lots of this antibody have been used on hamster, canine, and rat samples.
PA1-063A has been successfully used in Western blot procedures. By Western blot, this antibody detects an ~160 kDa protein representing EEA1 from mouse brain extract. Previous lots of this antibody have been used in immunoprecipitation and immunofluorescence procedures.
The PA1-063A immunogen is a synthetic peptide corresponding to residues T(1391) P S S K K P V R V C D A C F N D L Q G(1410) of human EEA1. The PA1-063A immunizing peptide (Cat. # PEP-079) is available for use in neutralization and control experiments.
Early endosomal antigen 1 (EEA1) is a 162 kDa membrane bound protein component specific to the early endosome and is essential for fusion between early endocytic vesicles. Early endosomes are cytoplasmic compartments which fuse with endocytic vesicles for redistribution of extracellular compounds to alternate destinations. Zinc-finger-like domains, reminiscent of those found in nucleic acid binding proteins, are located in the amino and carboxyl-terminal domains of EEA1. The carboxyl-terminal zinc-finger-like-domain is conserved in several other non-nuclear proteins, some of which are also involved in intracellular protein trafficking. In addition, this domain is an authentic zinc-binding domain and is critical to the intracellular localization of EEA1. Membrane association of EEA1 has been shown dependent on phosphatidyl 3-kinase activity, inhibitors of which cause EEA1 to dissociate from early endosomes.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
GM130 gain-of-function induces cell pathology in a model of lysosomal storage disease.
PA1-063A was used in immunocytochemistry to investigate the role of GM130 during lysosomal biogenesis
|Roy E,Bruyère J,Flamant P,Bigou S,Ausseil J,Vitry S,Heard JM||Human molecular genetics (21:1481)||2012|
Essential role of endocytosis of the type II transmembrane serine protease TMPRSS6 in regulating its functionality.
PA1-063A was used in immunocytochemistry to investigate the mechanism for TMPRSS6 function in iron homeostasis
|Béliveau F,Brulé C,Désilets A,Zimmerman B,Laporte SA,Lavoie CL,Leduc R||The Journal of biological chemistry (286:29035)||2011|
Assembly of connexin43 into gap junctions is regulated differentially by E-cadherin and N-cadherin in rat liver epithelial cells.
PA1-063A was used in immunocytochemistry to investigate the regulation of the Cx43 assembly by E-cadherin and N-cadherin
|Govindarajan R,Chakraborty S,Johnson KE,Falk MM,Wheelock MJ,Johnson KR,Mehta PP||Molecular biology of the cell (21:4089)||2010|
HIV-1 Nef binds a subpopulation of MHC-I throughout its trafficking itinerary and down-regulates MHC-I by perturbing both anterograde and retrograde trafficking.
PA1-063A was used in immunocytochemistry to study the mechanisms of HLA-1 down regulation by HIV-1 Nef.
|Yi L,Rosales T,Rose JJ,Chowdhury B,Chaudhury B,Knutson JR,Venkatesan S||The Journal of biological chemistry (285:30884)||2010|
E-cadherin differentially regulates the assembly of Connexin43 and Connexin32 into gap junctions in human squamous carcinoma cells.
PA1-063A was used in immunocytochemistry to investigate the effect of E-cadherin on gap junction formation in cancer cells
|Chakraborty S,Mitra S,Falk MM,Caplan SH,Wheelock MJ,Johnson KR,Mehta PP||The Journal of biological chemistry (285:10761)||2010|
Modulation of endocytic trafficking and apical stability of CFTR in primary human airway epithelial cultures.
PA1-063A was used in immunocytochemistry to investigate the trafficking and stability of CFTR in primary human airway epithelial cells
|Cholon DM,O'Neal WK,Randell SH,Riordan JR,Gentzsch M||American journal of physiology. Lung cellular and molecular physiology (298:L304)||2010|
Differential functions of ApoER2 and very low density lipoprotein receptor in Reelin signaling depend on differential sorting of the receptors.
PA1-063A was used in immunocytochemistry to investigate the different roles of ApoER2 and very low density lipoprotein receptor in Reelin signaling and their mechanism
|Duit S,Mayer H,Blake SM,Schneider WJ,Nimpf J||The Journal of biological chemistry (285:4896)||2010|
Misfolded BRICHOS SP-C mutant proteins induce apoptosis via caspase-4- and cytochrome c-related mechanisms.
PA1-063A was used in immunocytochemistry to study the mechanism that misfolded BRICHOS SP-C mutant proteins could induce apoptosis
|Mulugeta S,Maguire JA,Newitt JL,Russo SJ,Kotorashvili A,Beers MF||American journal of physiology. Lung cellular and molecular physiology (293:L720)||2007|
Urocortin trafficking in cerebral microvessel endothelial cells.
PA1-063A was used in immunocytochemistry to investigate the mechanism of urocortin endocytosis in cerebral microvessel endothelial cells
|Tu H,Kastin AJ,Bjorbaek C,Pan W||Journal of molecular neuroscience : MN (31:171)||2007|
UNC93B1 physically associates with human TLR8 and regulates TLR8-mediated signaling.
PA1-063A was used in immunohistochemistry to investigate the interaction between UNC93B1 and TLR8 and the regulation of TLR8-mediated signaling
|Itoh H,Tatematsu M,Watanabe A,Iwano K,Funami K,Seya T,Matsumoto M||PloS one (6:null)||2011|
Physiological relevance of cell-specific distribution patterns of CFTR, NKCC1, NBCe1, and NHE3 along the crypt-villus axis in the intestine.
PA1-063A was used in immunohistochemistry to investigate the distribution patterns of four ion transporters along the crypt-villus axis in the intestine
|Jakab RL,Collaco AM,Ameen NA||American journal of physiology. Gastrointestinal and liver physiology (300:G82)||2011|
The phospholipase A¿ enzyme complex PAFAH Ib mediates endosomal membrane tubule formation and trafficking.
PA1-063A was used in western blot to study the role of platelet-activating factor acetylhydrolase Ib (PAFAH) in endosomal export
|Bechler ME,Doody AM,Ha KD,Judson BL,Chen I,Brown WJ||Molecular biology of the cell (22:2348)||2011|
Abnormal trafficking and degradation of TLR4 underlie the elevated inflammatory response in cystic fibrosis.
PA1-063A was used in western blot to study the mechanism for the elevated inflammatory response in cystic fibrosis
|Bruscia EM,Zhang PX,Satoh A,Caputo C,Medzhitov R,Shenoy A,Egan ME,Krause DS||Journal of immunology (Baltimore, Md. : 1950) (186:6990)||2011|
Mitochondrially localized EGFR is independent of its endocytosis and associates with cell viability.
PA1-063A was used in western blot to study the localization and function of EGFR in mitochondria
|Yao Y,Wang G,Li Z,Yan B,Guo Y,Jiang X,Xi Z||Acta biochimica et biophysica Sinica (42:763)||2010|
Endocytosis in cultured neurons is altered by chronic alcohol exposure.
PA1-063A was used in western blot to investigate the influence of chronic alcohol consumption on neuronal endocytosis
|Marín MP,Esteban-Pretel G,Ponsoda X,Romero AM,Ballestín R,López C,Megías L,Timoneda J,Molowny A,Canales JJ,Renau-Piqueras J||Toxicological sciences : an official journal of the Society of Toxicology (115:202)||2010|