Overview

  • Product name

  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Serum 9 3.12%
    Inter-assay
    Sample n Mean SD CV%
    Serum 3 6.88%
  • Sample type

    Urine, Serum, Heparin Plasma, EDTA Plasma, Citrate Plasma
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    4.98 pg/ml
  • Range

    11.72 pg/ml - 750 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Urine 112 105% - 116%
    Serum 97 95% - 98%
    Heparin Plasma 100 97% - 101%
    EDTA Plasma 102 97% - 106%
    Citrate Plasma 104 99% - 108%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

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

    tPA in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of tPA protein in mouse and rat serum, plasma, and urine.


    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.


    Tissue plasminogen activator (tPA) is a serine protease belonging to the peptidase S1 family found on endothelial cells encoded by the gene Plat. tPA is expressed as a 526- amino acid glycosylated protein containing a fibronectin type-1 domain, a EGF-like domain, two Kringle domains, and a C-terminal Peptidase S1 domain. tPA can be cleaved into two catalytically active, disulfide-linked chains at Arg-308 by the proteins plasmin, tissue kallikrein, or factor Xa. The main function of tPA is the breakdown of blood clots by catalyzing the conversion of plasminogen to plasmin, which degrades the fibrin matrix of the clot. tPA also plays a role in cell migration, tissue remodeling, as well as proteolytic degradation of proteins in the brain extracellular matrix.

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Pre-coated microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Wash Buffer PT (ab206977) 1 x 20ml
    Antibody Diluent 5BR 1 x 6ml
    10X Mouse/Rat tPA Capture Antibody 1 x 600µl
    10X Mouse/Rat tPA Detector Antibody 1 x 600µl
    Mouse/Rat tPA Lyophilized Recombinant Protein 2 vials
    Plate Seals 1 unit
    Sample Diluent NS 1 x 12ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas

  • Function

    Converts the abundant, but inactive, zymogen plasminogen to plasmin by hydrolyzing a single Arg-Val bond in plasminogen. By controlling plasmin-mediated proteolysis, it plays an important role in tissue remodeling and degradation, in cell migration and many other physiopathological events. Play a direct role in facilitating neuronal migration.
  • Tissue specificity

    Synthesized in numerous tissues (including tumors) and secreted into most extracellular body fluids, such as plasma, uterine fluid, saliva, gingival crevicular fluid, tears, seminal fluid, and milk.
  • Involvement in disease

    Note=Increased activity of TPA results in increased fibrinolysis of fibrin blood clots that is associated with excessive bleeding. Defective release of TPA results in hypofibrinolysis that can lead to thrombosis or embolism.
  • Sequence similarities

    Belongs to the peptidase S1 family.
    Contains 1 EGF-like domain.
    Contains 1 fibronectin type-I domain.
    Contains 2 kringle domains.
    Contains 1 peptidase S1 domain.
  • Domain

    Both FN1 and one of the kringle domains are required for binding to fibrin.
    Both FN1 and EGF-like domains are important for binding to LRP1.
    The FN1 domain mediates binding to annexin A2.
    The second kringle domain is implicated in binding to cytokeratin-8 and to the endothelial cell surface binding site.
  • Post-translational
    modifications

    The single chain, almost fully active enzyme, can be further processed into a two-chain fully active form by a cleavage after Arg-310 catalyzed by plasmin, tissue kallikrein or factor Xa.
    Differential cell-specific N-linked glycosylation gives rise to two glycoforms, type I (glycosylated at Asn-219) and type II (not glycosylated at Asn-219). The single chain type I glycoform is less readily converted into the two-chain form by plasmin, and the two-chain type I glycoform has a lower activity than the two-chain type II glycoform in the presence of fibrin.
    N-glycosylation of Asn-152; the bound oligomannosidic glycan is involved in the interaction with the mannose receptor.
    Characterization of O-linked glycan was studied in Bowes melanoma cell line.
  • Cellular localization

    Secreted > extracellular space.
  • Information by UniProt
  • Alternative names

    • Alteplase
    • DKFZp686I03148
    • Plasminogen activator tissue
    • Plasminogen activator tissue type
    • PLAT
    • Reteplase
    • t PA
    • T Plasminogen Activator
    • t-PA
    • T-plasminogen activator
    • Tissue plasminogen activator (t PA)
    • Tissue type plasminogen activator
    • Tissue-type plasminogen activator chain B
    • tPA
    • TPA_HUMAN
    • TPA1
    see all
  • Database links

Associated products

Applications

Our Abpromise guarantee covers the use of ab233615 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 tPA standard curve was prepared as described in Section 10.

  • The concentrations of tPA were measured in duplicates, interpolated from the tPA standard curves and corrected for sample dilution. Undiluted samples are as follows: serum 50%, plasma (heparin) 12.5%, plasma (EDTA) 12.5%, plasma (citrate) 12.5%, and urine 25%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean tPA concentration in the neat samples was determined to be 540 pg/mL in serum, 2,567 pg/mL in plasma (heparin), 1,971 pg/mL in plasma (EDTA), 3,141 pg/mL in plasma (citrate), and 2,223 pg/mL in urine.

  • The concentrations of tPA were measured in duplicates, interpolated from the tPA standard curves and corrected for sample dilution. Undiluted samples are as follows: serum 25%, plasma (citrate) 12.5%, and urine 50%. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean tPA concentration in the neat samples was determined to be 1,657 pg/mL in serum, 5,687 pg/mL in plasma (citrate), and 388 pg/mL in urine.

Protocols

References

ab233615 has not yet been referenced specifically in any publications.

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