MABS1330
Anti-N1-Phosphohistidine (1-pHis) Antibody, clone SC1-1
clone SC1-1, from rabbit
Sign Into View Organizational & Contract Pricing
All Photos(1)
N1-Phosphohistidine
Recommended Products
biological source
rabbit
Quality Level
antibody form
purified antibody
antibody product type
primary antibodies
clone
SC1-1, monoclonal
species reactivity
E. coli, human
species reactivity (predicted by homology)
all
technique(s)
affinity chromatography: suitable
dot blot: suitable
immunocytochemistry: suitable
western blot: suitable
isotype
IgG
shipped in
wet ice
target post-translational modification
phosphorylation (N1-pHis)
General description
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 peptides containing stable phosphoryltriazolylalanine analogues of 1-pHis and 3-pHis (1-pTza and 3-pTza) 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.
Specificity
Selectively detects proteins with histidine(s) phosphorylated at N1 of the imidazole ring (1-pHis), but not 3-pHis.
Target modification is not species specific.
Immunogen
Epitope: N1-phosphohistidine (1-pHis)
KLH-conjugated library of random peptides containing non-hydrolyzable phosphohistidine analogue 1-pTza.
Application
Anti-N1-Phosphohistidine (1-pHis) antibody, clone SC1-1 is an isomer-specific monoclonal Ab to specifically detect histidine phosphorylated at position N1. This purified mAb is backed by published data demonstrating performance in Western Blots, immunofluorescence & immunoaffinity purification.
Dot Blot Analysis: A representative lot detected recombinant human NM23-H1/NME1 in vitro autophosphorylation on His118, as well as peptides containing 1-pTza, but not 3-pTza phosphohistidine analogue. Clone SC1-1 is most reactive toward 1-pTza peptides based on NM23-H1/NME1 1-pHis118 sequence, less reactive toward those based on histone H4 1-pHis18 or ACLY 1-pHis760 sequence, least reactive toward GNB1 1-pHis266-based or Kca3.1 1-pHis358-based sequence and not reactive toward phosphorylated tyrosine (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
Western Blotting Analysis: A representative lot detected heat-sensitive histidine N1-phosphorylation (1-pHis) in multiple cell lysates (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
Immunocytochemistry Analysis: A representative lot detected N1-phosphohistidine (1-pHis) immunoreactivity distinct from that of 3-pHis in 4% paraformaldehyde-fixed HeLa cells and murine bone marrow-derived macrophages by fluorescent immunocytochemistry. The 1-pHis immunoreactivity was found in regions surrounding acidic compartments, but not inside these compartments or nuclei (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
Immunoaffinity Purification: A representative lot was cross-linked to protein A resins for immunoaffinity purification of 1-pHis proteins from cell lysates prior to LC-MS/MS analysis (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
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 minutes 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 representative lot detected heat-sensitive histidine N1-phosphorylation (1-pHis) in multiple cell lysates (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
Immunocytochemistry Analysis: A representative lot detected N1-phosphohistidine (1-pHis) immunoreactivity distinct from that of 3-pHis in 4% paraformaldehyde-fixed HeLa cells and murine bone marrow-derived macrophages by fluorescent immunocytochemistry. The 1-pHis immunoreactivity was found in regions surrounding acidic compartments, but not inside these compartments or nuclei (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
Immunoaffinity Purification: A representative lot was cross-linked to protein A resins for immunoaffinity purification of 1-pHis proteins from cell lysates prior to LC-MS/MS analysis (Fuhs, S.R., et al. (2015). Cell. 162(1):198-210).
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 minutes 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.
Quality
Evaluated by Western Blotting of NME1 autophosphorylation reaction.
Western Blotting Analysis: 0.3 µg/mL of this antibody detected recombinant human NME1 (NM23-H1) with N1-phosphohistidine (1-pHis) in a 5 µg aliquot of autophosphorylation reaction.
Western Blotting Analysis: 0.3 µg/mL of this antibody detected recombinant human NME1 (NM23-H1) with N1-phosphohistidine (1-pHis) in a 5 µg aliquot of autophosphorylation reaction.
Target description
Variable depending on the histidine-phosphorylated proteins.
Physical form
Format: Purified
Other Notes
Concentration: Please refer to lot specific datasheet.
recommended
WGK
WGK 1
Certificates of Analysis (COA)
Search for Certificates of Analysis (COA) by entering the products Lot/Batch Number. Lot and Batch Numbers can be found on a product’s label following the words ‘Lot’ or ‘Batch’.
Already Own This Product?
Find documentation for the products that you have recently purchased in the Document Library.
LHPP suppresses gastric cancer progression via the PI3K/AKT/mTOR signaling pathway.
Journal of Cancer, 13, 3584-3592 (2022)
Methods in molecular biology (Clifton, N.J.), 2077, 181-191 (2019-11-11)
Immunoblotting is a ubiquitous immunological technique that aids in detecting and quantifying proteins (including those of lower abundance) and their posttranslational modifications such as phosphorylation, acetylation, ubiquitylation, and sumoylation. The technique involves electrophoretically separating proteins on an SDS-PAGE gel, transferring
Oncotarget, 9(12), 10185-10202 (2018-03-15)
The NM23/NME gene was identified as a metastasis suppressor. It's re-expression inhibited cancer cell motility and suppressed metastasis, without effecting primary tumor size in multiple model systems. The mechanisms of NME suppression of motility and metastasis are incompletely known. Of
Methods in molecular biology (Clifton, N.J.), 2077, 209-224 (2019-11-11)
Immunofluorescence (IF) takes advantage of biological and physical mechanisms to identify proteins in cell or tissue samples, exploiting the specificity of antibodies and stimulated fluorescence light emission. Here, we describe an immunofluorescence staining method for the identification of histidine phosphorylated
Nature communications, 11(1), 518-518 (2020-01-26)
Ethylene plays essential roles during adaptive responses to water-saturating environments in rice, but knowledge of its signaling mechanism remains limited. Here, through an analysis of a rice ethylene-response mutant mhz1, we show that MHZ1 positively modulates root ethylene responses. MHZ1
Our team of scientists has experience in all areas of research including Life Science, Material Science, Chemical Synthesis, Chromatography, Analytical and many others.
Contact Technical Service