Western blot analysis of Estrogen Receptor (ER) was performed by loading the indicated amounts of recombinant full-length human ER alpha (Product # A15674) and ER beta (Product # A15664) proteins, and 10ul of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-Glycine polyacrylamide gel. Proteins were transferred to a PVDF membrane (Product # 88518) using the G2 Blotter (Product # 62288), and blocked with 5% milk in TBST for at least 1 hour at room temperature. ER alpha was detected at ~66kD (left panel) using an ER monoclonal antibody (Product # MA5-12328) at a dilution of 1:200 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG secondary antibody (Product # 31430) at a dilution of 1:40,000. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080). The blot was then stripped with Restore Plus Western blot stripping buffer (Product # 46430) for at least 15 minutes at room temperature, re-blocked with 5% milk in TBST for at least 1 hour at room temperature, and re-probed with an ER beta polyclonal antibody (Product # PA1-313, right panel) at a dilution of 1:2000 overnight at 4C, followed by an HRP-conjugated goat anti-rabbit IgG secondary antibody (Product # 31460) at a dilution of 1:40,000. Chemiluminescent detection was again performed using SuperSignal West Pico (Product # 34080).
|Tested species reactivity||Bovine, Human, Mouse, Rabbit, Rat|
|Published species reactivity||Hamster, Bovine, Human, Mouse, Not Applicable|
|Host / Isotype||Mouse / IgG1, lambda|
|Immunogen||Purified, SDS-denatured calf uterus estrogen receptor (ER)|
|Storage buffer||PBS, pH 7.4, with 0.2% BSA|
|Contains||0.09% sodium azide|
|Storage Conditions||4° C|
|Tested Applications||Dilution *|
|Gel Shift (GS)||1mg/mL|
|Immunoprecipitation (IP)||2 µg/ml|
|Western Blot (WB)||1:100-1:400|
* 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.
MA5-12328 targets Estrogen Receptor in WB, GS, and IP applications and shows reactivity with bovine, human, mouse, rabbit, and rat samples. By Western blot, MA5-12328 detects recombinant ER alpha, but may not be suitable for detection of endogenous protein.
The MA5-12328 immunogen is purified, SDS-denatured calf uterus estrogen receptor (ER).
Estrogen Receptors (ER) are members of the steroid family of nuclear receptors. There are two different forms of the estrogen receptor, alpha and beta, encoded by separate genes (ESR1 and ESR2, respectively). Each protein contains distinct functional domains required for transcriptional activation, binding to estrogen response elements (ERE) in DNA, constitutive dimerization, binding to heat shock proteins, and ligand recognition. ER is a ligand-activated transcription factor, that when bound to estrogen hormone, induces a conformational change that allows dimerization and binding to EREs. When bound to DNA, ER can positively or negatively regulate gene transcription through the recruitment of coactivator or corepressor proteins. There are several different isoforms of both ER alpha and ER beta. ER is an important regulator of growth and differentiation in the mammary gland. The presence of ER in breast tumors indicates an increased likelihood of response to anti-estrogen (e.g. tamoxifen) therapy.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Characterization of estrogen response element binding proteins as biomarkers of breast cancer behavior.
MA5-12328 was used in EMSA to study the prognostic value of estrogen resonse element binding proteins in breast cancer
|Kruer TL,Cummins TD,Powell DW,Wittliff JL||Clinical biochemistry (46:1739)||2013|
Classification of breast cancer cells on the basis of a functional assay for estrogen receptor.
MA5-12328 was used in EMSA to study the classification of estrogen receptor-positive breast tumor cells on the basis of a functional estrogen receptor assay
|Biswas DK,Averboukh L,Sheng S,Martin K,Ewaniuk DS,Jawde TF,Wang F,Pardee AB||Molecular medicine (Cambridge, Mass.) (4:454)||1998|
|Not Applicable||Not Cited||
Targeting BIG3-PHB2 interaction to overcome tamoxifen resistance in breast cancer cells.
MA5-12328 was used in western blot to report that ERAP treatment suppresses tamoxifen resistance and enhances tamoxifen responsiveness in ERalpha-positive breast cancer cells
|Yoshimaru T,Komatsu M,Matsuo T,Chen YA,Murakami Y,Mizuguchi K,Mizohata E,Inoue T,Akiyama M,Yamaguchi R,Imoto S,Miyano S,Miyoshi Y,Sasa M,Nakamura Y,Katagiri T||Nature communications (4:null)||2013|
Expression and subcellular distribution of native estrogen receptor beta in murine C2C12 cells and skeletal muscle tissue.
MA5-12328 was used in western blot to study the expression and subcellular distribution of estrogen receptor beta in murine C2C12 cells and skeletal muscle tissue
|Milanesi L,Vasconsuelo A,de Boland AR,Boland R||Steroids (74:489)||2009|
Identification of estrogen receptor beta expression in Chinese hamster ovary (CHO) cells and comparison of estrogen-responsive gene transcription in cells adapted to serum-free media.
MA5-12328 was used in western blot to study estrogen receptor beta expression in Chinese hamster ovary cells
|Thomas PB,Risinger KE,Klinge CM||The Journal of steroid biochemistry and molecular biology (86:41)||2003|
Affinity chromatography of human estrogen receptor-alpha expressed in Saccharomyces cerevisiae. Combination of heparin- and 17beta-estradiol-affinity chromatography.
MA5-12328 was used in western blot to develop a method for the affinity purification of human estrogen receptor-alpha expressed in Saccharomyces cerevisiae
|Feng W,Graumann K,Hahn R,Jungbauer A||Journal of chromatography. A (852:161)||1999|
Identification of a variant estrogen receptor lacking exon 4 and its coexpression with wild-type estrogen receptor in ovarian carcinomas.
MA5-12328 was used in western blot to identify and study the expression of a variant estrogen receptor lacking exon 4 in ovarian carcinoma cells
|Park W,Choi JJ,Hwang ES,Lee JH||Clinical cancer research : an official journal of the American Association for Cancer Research (2:2029)||1996|
Location analysis for the estrogen receptor-alpha reveals binding to diverse ERE sequences and widespread binding within repetitive DNA elements.
MA5-12328 was used in ChIP assay to study the nucleotide sequence binding specificity of estrogen receptor-alpha
|Mason CE,Shu FJ,Wang C,Session RM,Kallen RG,Sidell N,Yu T,Liu MH,Cheung E,Kallen CB||Nucleic acids research (38:2355)||2010|
Leukemogenic MLL fusion proteins bind across a broad region of the Hox a9 locus, promoting transcription and multiple histone modifications.
MA5-12328 was used in ChIP assay to study the mechanisms by which leukemogenic MLL proteins induce multiple modifications of histones
|Milne TA,Martin ME,Brock HW,Slany RK,Hess JL||Cancer research (65:11367)||2005|
Nuclear receptor target gene discovery using high-throughput chromatin immunoprecipitation.
MA5-12328 was used in ChIP assay to identify nuclear receptor target genes by means of chromatin immunoprecipitation
|Laganière J,Deblois G,Giguère V||Methods in enzymology (364:339)||2003|
Repressive domain of unliganded human estrogen receptor alpha associates with Hsc70.
MA5-12328 was used in immunoprecipitation to study the molecular basis of how unliganded human ERalpha is maintained silent with respect to target gene promoters
|Ogawa S,Oishi H,Mezaki Y,Kouzu-Fujita M,Matsuyama R,Nakagomi M,Mori E,Murayama E,Nagasawa H,Kitagawa H,Yanagisawa J,Yano T,Kato S||Genes to cells : devoted to molecular and cellular mechanisms (10:1095)||2005|
The effects of estrogen, its antagonist ICI 182, 780, and interferon-tau on the expression of estrogen receptors and integrin alphaV beta 3 on cycle day 16 in bovine endometrium.
MA5-12328 was used in immunohistochemistry to study the effects of estrogen and IFN-tau on the expression of estrogen receptors and integrin alphaV beta 3 in bovine endometrium
|Kimmins S,Russell GL,Lim HC,Hall BK,MacLaren LA||Reproductive biology and endocrinology : RB and E (1:null)||2003|
Occurrence of estrogen receptor alpha in bovine placentomes throughout mid and late gestation and at parturition.
MA5-12328 was used in immunohistochemistry to study the immunohistochemical expression of estrogen receptor-alpha in bovine placentomes
|Schuler G,Wirth C,Teichmann U,Failing K,Leiser R,Thole H,Hoffmann B||Biology of reproduction (66:976)||2002|