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由来生物
rabbit
品質水準
抗体製品の状態
affinity isolated antibody
抗体製品タイプ
primary antibodies
クローン
polyclonal
精製方法
affinity chromatography
交差性
all, human, sheep, mouse
テクニック
ELISA: suitable
dot blot: suitable
immunoprecipitation (IP): suitable
western blot: suitable
輸送温度
ambient
ターゲットの翻訳後修飾
phosphorylation (pHis )
遺伝子情報
human ... PHPT1(29085)
詳細
Phosphorylation plays an important role in regulating protein activities and various cellular signaling events in cells. Limited by the tools available for phosphohistidine (pHis) detection, the majority of studies focus on serine, threonine, and tyrosine phosphorylations. Histidine phosphorylation can occur at either N1 (1-pHis) or N3 (3-pHis) of the imidazole ring. The development of stable phosphoryltriazolylalanine analogues of pHis (pTza and pPza) allows the generation of antibodies for studying both histidine N1 and N3 phosphorylations in signaling events. There is growing evidence implicating His kinases in cancer and tumor metastasis and the first metastasis suppressor gene identified is one of the two known mammalian His kinases, Nm23-H1 (also known as NME1, nucleoside diphosphate kinase, or NDPK-A). Nm23-H1/NME1 and the closely related Nm23-H2 (NME2/NDPK-B) catalyze the transfer of phosphate from ATP onto Nucleoside-diphosphates (NDPs) through a 1-pHis enzyme intermediate. Nm23-H1/-H2 also possess His kinase activity, transferring the phosphate from the active site pHis onto a His in a target protein. Metabolic enzymes such as phosphoglycerate mutase (PGAM), succinyl CoA synthase (SCS), and ATP citrate lyase (ACL) also use pHis as an enzyme intermediate. Unlike NME1/2, PGAM uses 3-pHis as an enzyme intermediate. In addition to eukaryotes, histidine phosphorylation is well documented in bacterial two-component signaling pathways involved in chemotaxis, although the phosphate is transferred from the pHis formed in the receptor/sensor protein to Asp residues of an acceptor response regulator protein, and the receptor/sensor protein essentially functions as an aspartate kinase.
特異性
Target modification is not species-specific.
This rabbit polyclonal antibody reacted with pHis- and pPza-, but not His-, pTyr-, pSer-, or pThr-, conjugated BSA. Specificity testing using N1-pHis analog showed minimal cross-reactivity.
免疫原
KLH-conjugated non-hydrolyzable phosphohistidine analogue 4-phosphopyrazol-2-yl alanine (pPza).
アプリケーション
Research Category
細胞シグナル伝達
細胞シグナル伝達
Detect histidine phosphorylation using this rabbit polyclonal Anti-Phosphohistidine (pHis) , Cat. No. ABS1670, validated for use in Dot Blot, ELISA, Immunoprecipitation, and Western Blotting.
Dot Blot Analysis: A 1:1,200 dilution from a representative lot detected 5 µg of pHis- and pPza-, but not His-, pTyr-, pSer-, or pThr-, conjugated BSA (Courtesy of Bezaleel Mambwe, Richmond Muimo and RFW Jackson, Department of Infection, Immunity and Cardiovascular Disease/ Department of Chemistry, University of Sheffield, UK).
Western Blotting Analysis: A 1:120 dilution from a representative lot detected proteins with histidine phosphorylation in sheep trachea cytosolic extract and human cell lysates, including 16HBE14o-, HEK293T, THP-1, and THP-1-derived macrophages (Courtesy of Bezaleel Mambwe, Richmond Muimo and RFW Jackson, Department of Infection, Immunity and Cardiovascular Disease/ Department of Chemistry, University of Sheffield, UK).
ELISA Analysis: A representative lot detected pHis- and pPza-, but not pTyr-, pSer-, pThr-, conjugated BSA or unconjugated BSA (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Immunoprecipitation Analysis: A representative lot immunoprecipitated proteins with histidine phosphorylation from ovine airway epithelia extract (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Western Blotting Analysis: A representative lot detected histidine phosphorylated proteins in 16HBE14o- human bronchial epithelial cell lysate and in ovine airway epithelia extract. Acid (0.1 M HCl or 0.4 M acetic acid/0.1 M hydroxylamine), but not alkaline (0.1 M NaOH), treatment of the lysates abolished targets bands detection (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Western Blotting Analysis: A representative lot detected histidine phosphorylation of immunoprecipitated NDPK-A/B from ovine airway epithelia extract, as well as G -R/M from 16HBE14o- human bronchial epithelial cell lysate (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Note: DO NOT HEAT SAMPLES prior to phosphohistidine detection. Histidine phosphorylation is heat and acid labile. To generate negative control for specificity test, an aliquot of sample can be heated at 95ºC for 10-15 minutes to reverse histidine phosphorylation. Alternatively, an aliquot of sample can be incubated under acidified pH at 37ºC for 15 minunites to reduce histidine phosphorylation. Acidify each 100 µL sample with 25 µL of 1 M HCl before the incubation, then neutralize with 25 µL of 1 M NaOH prior to phosphohistidine detection.
Western Blotting Analysis: A 1:120 dilution from a representative lot detected proteins with histidine phosphorylation in sheep trachea cytosolic extract and human cell lysates, including 16HBE14o-, HEK293T, THP-1, and THP-1-derived macrophages (Courtesy of Bezaleel Mambwe, Richmond Muimo and RFW Jackson, Department of Infection, Immunity and Cardiovascular Disease/ Department of Chemistry, University of Sheffield, UK).
ELISA Analysis: A representative lot detected pHis- and pPza-, but not pTyr-, pSer-, pThr-, conjugated BSA or unconjugated BSA (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Immunoprecipitation Analysis: A representative lot immunoprecipitated proteins with histidine phosphorylation from ovine airway epithelia extract (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Western Blotting Analysis: A representative lot detected histidine phosphorylated proteins in 16HBE14o- human bronchial epithelial cell lysate and in ovine airway epithelia extract. Acid (0.1 M HCl or 0.4 M acetic acid/0.1 M hydroxylamine), but not alkaline (0.1 M NaOH), treatment of the lysates abolished targets bands detection (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Western Blotting Analysis: A representative lot detected histidine phosphorylation of immunoprecipitated NDPK-A/B from ovine airway epithelia extract, as well as G -R/M from 16HBE14o- human bronchial epithelial cell lysate (Lilley, M., et al. (2015). Chem. Commun. (Camb). 51(34):7305-7308).
Note: DO NOT HEAT SAMPLES prior to phosphohistidine detection. Histidine phosphorylation is heat and acid labile. To generate negative control for specificity test, an aliquot of sample can be heated at 95ºC for 10-15 minutes to reverse histidine phosphorylation. Alternatively, an aliquot of sample can be incubated under acidified pH at 37ºC for 15 minunites to reduce histidine phosphorylation. Acidify each 100 µL sample with 25 µL of 1 M HCl before the incubation, then neutralize with 25 µL of 1 M NaOH prior to phosphohistidine detection.
品質
Evaluated by Western Blotting in HEK293 cell lysate.
Western Blotting Analysis: A 1:200 dilution of this antibody detected histidine-phosphorylated proteins in 10 µg of HEK293 cell lysate.
Western Blotting Analysis: A 1:200 dilution of this antibody detected histidine-phosphorylated proteins in 10 µg of HEK293 cell lysate.
ターゲットの説明
Variable depending on the histidine-phosphorylated proteins.
物理的形状
Affinity purified.
Purified rabbit polyclonal antibody in buffer containing 100 mM glycine pH 2.2, 100 mM Tris pH 10 with 0.05% sodium azide.
保管および安定性
Stable for 1 year at 2-8°C from date of receipt.
その他情報
Concentration: Please refer to lot specific datasheet.
免責事項
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
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保管分類コード
12 - Non Combustible Liquids
WGK
WGK 1
引火点(°F)
Not applicable
引火点(℃)
Not applicable
適用法令
試験研究用途を考慮した関連法令を主に挙げております。化学物質以外については、一部の情報のみ提供しています。 製品を安全かつ合法的に使用することは、使用者の義務です。最新情報により修正される場合があります。WEBの反映には時間を要することがあるため、適宜SDSをご参照ください。
Jan Code
ABS1670:
試験成績書（COA）
製品のロット番号・バッチ番号を入力して、試験成績書（COA） を検索できます。ロット番号・バッチ番号は、製品ラベルに「Lot」または「Batch」に続いて記載されています。
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