Phospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA Kit (ab115343)

Overview

  • Product name
    Phospho S232 PDH E1 alpha protein (PDHA1) Profiling ELISA Kit
    See all Pyruvate Dehydrogenase E1-alpha subunit kits
  • Detection method
    Colorimetric
  • Precision
    Intra-assay
    Sample n Mean SD CV%
    1 8 = 8.3%
    Inter-assay
    Sample n Mean SD CV%
    1 4 = 7.7%
  • Sample type
    Cell culture extracts, Tissue Extracts
  • Assay type
    Sandwich (quantitative)
  • Sensitivity
    = 31 µg/ml
  • Range
    31 µg/ml - 1000 µg/ml
  • Assay duration
    Multiple steps standard assay
  • Species reactivity
    Reacts 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.

  • Notes

    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 applications
    Suitable for: ELISAmore details
  • Platform
    Microplate

Properties

  • Storage instructions
    Store 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
    HRP Development Solution 1 x 12ml
  • Research areas
  • Function
    The 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 specificity
    Ubiquitous.
  • Involvement in disease
    Defects 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.
  • Post-translational
    modifications
    Phosphorylation at Ser-293 by PDK family kinases blocks the access to active site, and inactivates the enzyme.
  • Cellular localization
    Mitochondrion matrix.
  • Information by UniProt
  • Alternative names
    • mitochondrial
    • ODPA_HUMAN
    • PDHA1
    • PDHCE1A
    • PDHE1-A type I
    • PHE1A
    • 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
    see all
  • Database links

Applications

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.

Application Abreviews Notes
ELISA Use at an assay dependent concentration.

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.

Protocols

References

ab115343 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

Unfortunately we are not able to sell the detection antibodies from this kit individually.

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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