Overview

  • Product name

    Ras GTPase ELISA Kit (Chemiluminescent)
  • Detection method

    Luminescent
  • Sample type

    Cell culture extracts
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    > 3000 ng/well
  • Range

    3000 ng/well - 25000 ng/well
  • Assay time

    4h 30m
  • Assay duration

    Multiple steps standard assay
  • Species reactivity

    Reacts with: Mouse, Human
  • Product overview

    ab134640 Ras GTPase ELISA Kit is designed specifically for the study of Ras activation and can be used to study novel signaling pathways for activating Ras. The kit can also be used as a diagnostic test to detect oncogenic Ras related to malignancy. Ras GTPase ELISA Kits contain a Raf-RBD protein fused to GST that will be coated onto the provided 96-well (12 x 8 well strips), glutathione-coated plate. The activated Ras contained in cellular extract specifically binds to Raf-RBD, while inactive Ras does not bind. Bound Ras is detected by incubating with a primary antibody that detects H-Ras in mouse and H- & K-Ras in human samples. Addition of a secondary antibody conjugated to horseradish peroxidase (HRP) and developing solution provides a sensitive chemiluminescent readout that is easily quantified by luminescence. The 96-well plate with individual strips of 12 wells is suitable for manual use or high-throughput screening applications.

     

    Ras GTPase ELISA Kit specifically detects activated H- and K-Ras in Human and H-Ras in rodent samples.

  • Tested applications

    Suitable for: ELISAmore details
  • Platform

    Microplate

Properties

  • Storage instructions

    Please refer to protocols.
  • Components 1 x 96 tests
    10X Antibody Binding Buffer 1 x 2.2ml
    10X Wash Buffer 2 x 22ml
    96-well assay plate 1 unit
    Anti-rat HRP-conjugated IgG 1 x 11µl
    Chemiluminescent Reagent 1 x 2ml
    GST-Raf-RBD 4 x 25µl
    HeLa whole-cell extract (EGF treated) 2 x 40µl
    H-Ras antibody 1 x 11µl
    Lysis Buffer 1 x 50ml
    Plate sealer 1 unit
    Protease Inhibitor Cocktail 1 x 500µl
    Reaction Buffer 1 x 4ml
  • Research areas

  • Function

    Ras proteins bind GDP/GTP and possess intrinsic GTPase activity.
  • Involvement in disease

    Defects in HRAS are the cause of faciocutaneoskeletal syndrome (FCSS) [MIM:218040]. A rare condition characterized by prenatally increased growth, postnatal growth deficiency, mental retardation, distinctive facial appearance, cardiovascular abnormalities (typically pulmonic stenosis, hypertrophic cardiomyopathy and/or atrial tachycardia), tumor predisposition, skin and musculoskeletal abnormalities.
    Defects in HRAS are the cause of congenital myopathy with excess of muscle spindles (CMEMS) [MIM:218040]. CMEMS is a variant of Costello syndrome.
    Defects in HRAS may be a cause of susceptibility to Hurthle cell thyroid carcinoma (HCTC) [MIM:607464]. Hurthle cell thyroid carcinoma accounts for approximately 3% of all thyroid cancers. Although they are classified as variants of follicular neoplasms, they are more often multifocal and somewhat more aggressive and are less likely to take up iodine than are other follicular neoplasms.
    Note=Mutations which change positions 12, 13 or 61 activate the potential of HRAS to transform cultured cells and are implicated in a variety of human tumors.
    Defects in HRAS are a cause of susceptibility to bladder cancer (BLC) [MIM:109800]. A malignancy originating in tissues of the urinary bladder. It often presents with multiple tumors appearing at different times and at different sites in the bladder. Most bladder cancers are transitional cell carcinomas. They begin in cells that normally make up the inner lining of the bladder. Other types of bladder cancer include squamous cell carcinoma (cancer that begins in thin, flat cells) and adenocarcinoma (cancer that begins in cells that make and release mucus and other fluids). Bladder cancer is a complex disorder with both genetic and environmental influences.
    Note=Defects in HRAS are the cause of oral squamous cell carcinoma (OSCC).
  • Sequence similarities

    Belongs to the small GTPase superfamily. Ras family.
  • Post-translational
    modifications

    Palmitoylated by the ZDHHC9-GOLGA7 complex. A continuous cycle of de- and re-palmitoylation regulates rapid exchange between plasma membrane and Golgi.
    S-nitrosylated; critical for redox regulation. Important for stimulating guanine nucleotide exchange. No structural perturbation on nitrosylation.
  • Cellular localization

    Cell membrane. Golgi apparatus membrane. The active GTP-bound form is localized most strongly to membranes than the inactive GDP-bound form (By similarity). Shuttles between the plasma membrane and the Golgi apparatus.
  • Information by UniProt
  • Alternative names

    • C-BAS/HAS
    • c-H-ras
    • C-HA-RAS1
    • CTLO
    • GTPase HRas
    • GTPase KRas
    • GTPase NRas
    • H-Ras-1
    • H-RASIDX
    • Ha-Ras
    • HAMSV
    • HRAS
    • HRAS1
    • K RAS2A
    • K RAS2B
    • K RAS4A
    • K RAS4B
    • K-RAS
    • KRAS
    • KRAS1
    • KRAS2
    • N-RAS
    • N-terminally processed
    • NRAS
    • NRAS1
    • p21ras
    • RASH_HUMAN
    • RASH1
    • RASK2
    • Transforming protein p21
    • v Ha ras Harvey rat sarcoma viral oncogene homolog
    • v Ki ras2 Kirsten rat sarcoma viral oncogene homolog
    • v ras neuroblastoma RAS viral oncogene homolog
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab134640 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

  • Quantification of activated Ras: Increasing amounts of whole-cell extracts from unstimulated 293T/17 and EGF stimulated HeLa cells were assayed for Ras activity using ab134640 Ras GTPase ELISA Kit. To illustrate the Kit’s specificity for activated Ras, 293T/17 cells which do not contain basal levels of activated Ras were used as a negative control. Data was also shown for unstimulated HeLa cells, which do contain basal levels of activated Ras. This data is provided for demonstration only.

Protocols

References

This product has been referenced in:

  • Yeung Y  et al. K-Ras mutation and amplification status is predictive of resistance and high basal pAKT is predictive of sensitivity to everolimus in biliary tract cancer cell lines. Mol Oncol 11:1130-1142 (2017). Read more (PubMed: 28544747) »
  • Feng C  et al. miRNA-556-3p promotes human bladder cancer proliferation, migration and invasion by negatively regulating DAB2IP expression. Int J Oncol 50:2101-2112 (2017). Read more (PubMed: 28440444) »
See all 2 Publications for this product

Customer reviews and Q&As

Answer


You will need to check with the manufacturer / manual of your plate reader to make sure it is capable of reading chemiluminescence.

Chemiluminescence is most commonly measured using a luminometer although some plate readers have an option to read chemiluminescence or can be adapted to measure total light output. If a wavelength must be selected, measurement at 425nm gives a crude indication of chemiluminescence. Some fluorescent plate readers can also be used if excitation is turned off.

Read More

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