Phospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA Kit (ab115343)
- Product namePhospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA KitSee all Pyruvate Dehydrogenase E1-alpha subunit kits ...
- Detection methodColorimetric
Intra-assay Sample n Mean SD CV% 1 8 = 8.3% Inter-assay Sample n Mean SD CV% 1 4 = 7.7%
- Tests1 x 96 test
- Sample typeCell culture extracts, Tissue Extracts
- Assay typeSandwich (quantitative)
- Sensitivity= 31 µg/ml
- Range31 µg/ml - 1000 µg/ml
- Assay durationMultiple steps standard assay
- Species reactivityReacts with: Mouse, Rat, Cow, Human
- Product overview
ab115343 is an in vitro enzyme-linked immunosorbent assay to determine the levels of phospho S232 PDHA1 protein in cell and tissue lysates. The assay employs a mouse antibody specific for PDHA1 protein coated on a 96-well plate. Samples are pipetted into the wells and PDHA1 protein present in the sample is bound to the wells by the immobilized antibody. The wells are washed and a rabbit anti-phospho S232 PDHA1 protein detector antibody is added. After washing away unbound detector antibody, HRP-conjugated anti-rabbit antibody is pipetted into the wells. The wells are again washed, an HRP substrate solution (TMB) is added to the wells and color develops in proportion to the amount of phospho S232 PDHA1 protein bound. The developing blue color is measured at 600 nm. Optionally the reaction can be stopped by adding hydrochloric acid which changes the color from blue to yellow and the intensity can be measured at 450 nm.
Store all components at 4°C. The kits are stable for at least 6
months. Unused microplate strips should be returned to the pouch
containing the desiccant and resealed.
- Tested applicationsELISA more details
- Storage instructionsStore at +4°C. Please refer to protocols.
Components 1 x 96 tests 10X Blocking Buffer 1 x 6ml 10X HRP Label 1 x 1ml 10X phospho S232 PDHA1 protein Detector Antibody 1 x 0.7ml 20X Buffer 1 x 20ml Extraction Buffer 1 x 15ml Microplate 96 antibody coated wells in 12 strips 1 unit TMB Development Solution 1 x 12ml
- FunctionThe pyruvate dehydrogenase complex catalyzes the overall conversion of pyruvate to acetyl-CoA and CO(2). It contains multiple copies of three enzymatic components: pyruvate dehydrogenase (E1), dihydrolipoamide acetyltransferase (E2) and lipoamide dehydrogenase (E3).
- Tissue specificityUbiquitous.
- Involvement in diseaseDefects in PDHA1 are a cause of pyruvate dehydrogenase E1-alpha deficiency (PDHAD) [MIM:312170]. An enzymatic defect causing primary lactic acidosis in children. It is associated with a broad clinical spectrum ranging from fatal lactic acidosis in the newborn to chronic neurologic dysfunction with structural abnormalities in the central nervous system without systemic acidosis.
Defects in PDHA1 are the cause of X-linked Leigh syndrome (X-LS) [MIM:308930]. X-LS is an early-onset progressive neurodegenerative disorder with a characteristic neuropathology consisting of focal, bilateral lesions in one or more areas of the central nervous system, including the brainstem, thalamus, basal ganglia, cerebellum, and spinal cord. The lesions are areas of demyelination, gliosis, necrosis, spongiosis, or capillary proliferation. Clinical symptoms depend on which areas of the central nervous system are involved. The most common underlying cause is a defect in oxidative phosphorylation. LS may be a feature of a deficiency of any of the mitochondrial respiratory chain complexes.
modificationsPhosphorylation at Ser-293 by PDK family kinases blocks the access to active site, and inactivates the enzyme.
- Cellular localizationMitochondrion matrix.
- PDHE1-A type I
- Pyruvate dehydrogenase (lipoamide) alpha 1
- Pyruvate dehydrogenase complex E1 alpha polypeptide 1
- Pyruvate dehydrogenase E1 component subunit alpha
- Pyruvate dehydrogenase E1 component subunit alpha somatic form mitochondrial
- Pyruvate dehydrogenase, alpha-1
- somatic form
Our Abpromise guarantee covers the use of ab115343 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Phospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA Kit images
The PDHA1 bound from undosed HeLa cells was subject to in-well kinase treatment (PDK1&3) or in-well phosphatase treatment (PDP1) according to the supplementary protocol shown below. Untreated cells showed a significant endogenous phosphorylation signal at S232 which is only slightly increased by kinase treatment. Conversely phosphatase treatment was able to significantly reduce the phospho S232 signal from the endogenous levels.
HeLa cells were cultured for 4 hours in media supplemented with DCA (20mM) to specifically inhibit mitochondrial PDH kinases, or NaF (20mM), a general inhibitor of serine/threonine protein phosphatases. The DCA treatment reduced the level of phospho S232. Conversely NaF treatment, to inhibit cellular serine phosphatases, increased the phosphorylation level of S232.
Example control sample curve for HepG2 cell extract. The phosphorylation state of PDHA1 can vary by treatment but also by cell culture conditions such as media supplements, nutrients and also cell density.
Example control sample curve for HeLa cell extract. The phosphorylation state of PDHA1 can vary by treatment but also by cell culture conditions such as media supplements, nutrients and also cell density.
References for Phospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA Kit (ab115343)
ab115343 has not yet been referenced specifically in any publications.