Overview

  • Product name

    Human Midkine ELISA Kit
    See all Midkine kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    HepG2 8 2%
    Inter-assay
    Sample n Mean SD CV%
    HepG2 3 3.9%
  • Sample type

    Cell culture supernatant, Serum, Cell culture extracts, Heparin Plasma, EDTA Plasma, Citrate Plasma
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    14.4 pg/ml
  • Range

    39.1 pg/ml - 2500 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture supernatant 100 82% - 115%
    Serum 114 113% - 115%
    Cell culture extracts 156 152% - 160%
    Heparin Plasma 83 73% - 90%
    EDTA Plasma 112 109% - 114%
    Citrate Plasma 128 115% - 137%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Dog, Human
    Does not react with: Mouse, Rat, Cow, Pig
  • Product overview

    As of August 14, 2019, Human Midkine SimpleStep ELISA® kit has been re-developed. We have identified new recombinant monoclonal antibodies to provide improved performance and consistency when quantifying Midkine protein in human serum, plasma, cell culture supernatant, cell and tissue extracts.


    The SimpleStep ELISA™ employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.

  • Notes

    Midkine (MK), also known as neurite growth-promoting factor 2 (NEGF2), is a 13kDa heparin-binding growth factor or cytokine composed of two domains: The N-terminally located N-domain and the C-terminally located C-domain, which are connected by a hinge. The C-domain plays a role in neuronal development, whereas the N-domain is important for protein stability and dimerization. MK is involved in development, reproduction, repair, inflammation, innate immunity, control of blood pressure and angiogenesis.

    MK is strongly expressed during embryogenesis and due to its distribution in the embryo it has been proposed to play a role in neurogenesis, epithelial-mesenchymal interactions and mesoderm remodeling. Expression in adult tissues is restricted to the kidney, epidermis, bronchial epithelium, lymphocytes and macrophages, but it is strongly expressed in the brain, kidney, blood vessels and heart after tissue injury as well as during inflammation, infection and oncogenesis. During inflammation, substratum-bound MK enhances neutrophil and macrophage migration directly and through induction of chemokine expression. On the other hand, soluble MK is associated with differentiation of regulatory T-cells, induction of epithelial-mesenchymal transition, angiogenesis, fibrinolytic and anti-microbial activity.

    MK signaling is mediated by cell surface receptors as well as membrane proteins such as Protein Tyrosine Phosphatase ζ (PTPζ), low density lipoprotein receptor-related proteins (LRP), Notch2, integrins, anaplastic lymphoma kinase (ALK) and neuroglycan C. Binding of MK to PTPζ induces tyrosine phosphorylation in β-catenin and Wnt signaling inhibition. Furthermore it induces phosphorylation of PI3K, MAPK, PKC and Src family kinase. Binding of MK to LRP leads to embryonic neuronal survival and prevention of hypoxic injury via Akt and HIF1α. Binding of MK to integrins activates focal adhesion kinase, paxillin and STAT1α pathway leading to increase invasiveness of cancer cells. Activation of ALK by MK leads to phosphorylation of IRS-1, MAPK, PI3K and activation of NF-κB.

    Due to its multifunctionality, MK has become an emerging target of drug development for the treatment of multiple diseases. On the one hand, administration of MK ameliorates ischemic injury, enhances oocyte maturation and promotes neurogenesis therefore limiting the progression of neurodegenerative diseases. However on the other hand, due to its over-expression in malignant tumors and in inflammation, MK inhibitors may be useful in the treatment of cancer, multiple sclerosis, hypertension and osteoporosis.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Human Midkine Capture Antibody 1 x 600µl
    10X Human Midkine Detector Antibody 1 x 600µl
    10X Wash Buffer PT (ab206977) 1 x 20ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    Antibody Diluent CPI - HAMA Blocker (ab193969) 1 x 6ml
    Human Midkine Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS (ab193972) 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Function

    Developmentally regulated, secreted growth factor homologous to pleiotrophin (PTN), which has heparin binding activity. Binds anaplastic lymphoma kinase (ALK) which induces ALK activation and subsequent phosphorylation of the insulin receptor substrate (IRS1), followed by the activation of mitogen-activated protein kinase (MAPK) and PI3-kinase, and the induction of cell proliferation. Involved in neointima formation after arterial injury, possibly by mediating leukocyte recruitment. Also involved in early fetal adrenal gland development.
  • Tissue specificity

    Expressed in various tumor cell lines. In insulinoma tissue predominantly expressed in precancerous lesions.
  • Sequence similarities

    Belongs to the pleiotrophin family.
  • Cellular localization

    Secreted.
  • Information by UniProt
  • Alternative names

    • Amphiregulin-associated protein
    • ARAP
    • Mdk
    • Midgestation and kidney protein
    • Midkine
    • MK
    • MK_HUMAN
    • Neurite outgrowth-promoting factor 2
    • Neurite outgrowth-promoting protein
    see all
  • Database links

Associated products

Applications

Our Abpromise guarantee covers the use of ab193761 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
Sandwich ELISA Use at an assay dependent concentration.

Images

  • SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

     

  • The Midkine standard curve was prepared as described in Section 10. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.

  • The concentrations of Midkine were measured in duplicates, interpolated from the Midkine standard curves and corrected for sample dilution. Undiluted samples are as follows: HepG2 cell culture supernatant 2.5%.  The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Midkine concentration was determined to be 6,242 pg/mL in serum.

  • The concentrations of Midkine were measured in duplicate and interpolated from the Midkine standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Midkine concentration was determined to be 2641 pg/mL in HepG2 cell extract and 1179 pg/mL in SH-SY5Y cell extract.

  • The concentrations of Midkine were measured in duplicates, interpolated from the Midkine standard curves and corrected for sample dilution. Undiluted samples are as follows: serum 50%, plasma (EDTA) 50%, plasma (citrate) 25%, and plasma (heparin) 50%.  The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Midkine concentration was determined to be 243 pg/mL in serum, 183 pg/mL in plasma (EDTA), 449 pg/mL plasma (citrate) and 303 pg/mL in plasma (heparin).

  • Interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean Midkine concentration was determined to be 164 pg/mL with a range of 69 – 594 pg/mL.

Protocols

References

This product has been referenced in:

  • Meng X  et al. DNA damage repair alterations modulate M2 polarization of microglia to remodel the tumor microenvironment via the p53-mediated MDK expression in glioma. EBioMedicine 41:185-199 (2019). Read more (PubMed: 30773478) »
  • Rice LM  et al. Serum biomarker for diagnostic evaluation of pulmonary arterial hypertension in systemic sclerosis. Arthritis Res Ther 20:185 (2018). Read more (PubMed: 30115106) »
See all 3 Publications for this product

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