Overview

  • Product name

    Bradykinin ELISA kit
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Buffer 20 695.4pg/ml 4.6%
    Buffer 20 208.7pg/ml 6.2%
    Buffer 20 73.7pg/ml 9.9%
    Inter-assay
    Sample n Mean SD CV%
    Buffer 700.4pg/ml = 15%
    Buffer 209.3pg/ml = 10.3%
    Buffer 66.1pg/ml = 11.9%
  • Sample type

    Urine, Serum, Plasma
  • Assay type

    Competitive
  • Sensitivity

    24.8 pg/ml
  • Range

    11.7 pg/ml - 30000 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Urine 109 10pg/ml - 2000pg/ml
    Serum 113 100pg/ml - 2000pg/ml
    Plasma 110 100pg/ml - 20000pg/ml

  • Assay time

    3h 00m
  • Assay duration

    Multiple steps standard assay
  • Species reactivity

    Reacts with: Species independent
  • Product overview

    Bradykinin ELISA kit is a competitive Enzyme-Linked Immunosorbent Assay designed for the accurate quantitative measurement of Bradykinin in plasma, serum and urine.


    A goat anti-Rabbit IgG antibody has been precoated onto 96-well plates. Standards or test samples are added to the wells, along with a solution of Bradykinin conjugated to biotin, followed by a solution of polyclonal antibody to Bradykinin. The plate is washed to remove unbound reagents. A solution of streptavidin-HRP conjugate is then added. After further incubation the excess reagents are washed away and TMB substrate is added, which is catalyzed by HRP to generate a yellow color. A stop solution changes this color from yellow to blue, and the intensity of this blue coloration is inversely proportional to the amount of Bradykinin captured in the plate.

  • Tested applications

    Suitable for: Competitive ELISAmore details
  • Platform

    Microplate

Properties

  • Storage instructions

    Please refer to protocols.
  • Components 1 x 96 tests
    20X Wash Buffer Concentrate 1 x 27ml
    Assay Buffer 16 1 x 55ml
    Bradykinin Antibody 1 x 5ml
    Bradykinin Conjugate 1 x 5ml
    Bradykinin Standard 2 vials
    Goat anti-rabbit IgG Microplate (12 x 8 wells) 1 unit
    HRP- Streptavidin Conjugate 1 x 12.5µg
    Plate Sealer 2 units
    Stop Solution 2 1 x 10ml
    TMB Substrate 2 x 10ml
  • Research areas

  • Function

    (1) Kininogens are inhibitors of thiol proteases; (2) HMW-kininogen plays an important role in blood coagulation by helping to position optimally prekallikrein and factor XI next to factor XII; (3) HMW-kininogen inhibits the thrombin- and plasmin-induced aggregation of thrombocytes; (4) the active peptide bradykinin that is released from HMW-kininogen shows a variety of physiological effects: (4A) influence in smooth muscle contraction, (4B) induction of hypotension, (4C) natriuresis and diuresis, (4D) decrease in blood glucose level, (4E) it is a mediator of inflammation and causes (4E1) increase in vascular permeability, (4E2) stimulation of nociceptors (4E3) release of other mediators of inflammation (e.g. prostaglandins), (4F) it has a cardioprotective effect (directly via bradykinin action, indirectly via endothelium-derived relaxing factor action); (5) LMW-kininogen inhibits the aggregation of thrombocytes; (6) LMW-kininogen is in contrast to HMW-kininogen not involved in blood clotting.
  • Tissue specificity

    Secreted in plasma. T-kinin is detected in malignant ovarian, colon and breast carcinomas, but not in benign tumors.
  • Involvement in disease

    Defects in KNG1 are the cause of high molecular weight kininogen deficiency (HMWK deficiency) [MIM:228960]. HMWK deficiency is an autosomal recessive coagulation defect. Patients with HWMK deficiency do not have a hemorrhagic tendency, but they exhibit abnormal surface-mediated activation of fibrinolysis.
  • Sequence similarities

    Contains 3 cystatin domains.
  • Post-translational
    modifications

    Bradykinin is released from kininogen by plasma kallikrein.
    Hydroxylation of Pro-383 occurs prior to the release of bradykinin.
    Phosphorylation sites are present in the extracelllular medium.
    N- and O-glycosylated. O-glycosylated with core 1 or possibly core 8 glycans.
  • Cellular localization

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

    • Alpha-2-thiol proteinase inhibitor
    • BDK
    • Fitzgerald factor
    • High molecular weight kininogen
    • HMWK
    • Ile-Ser-Bradykinin
    • Kallidin I
    • Kallidin II
    • Kininogen 1
    • KNG
    • KNG1
    • KNG1_HUMAN
    • Low molecular weight growth-promoting factor
    • Williams-Fitzgerald-Flaujeac factor
    see all

Applications

Our Abpromise guarantee covers the use of ab136936 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
Competitive ELISA Use at an assay dependent concentration.

Images

  • Representative standard curve using ab136936.

Protocols

References

This product has been referenced in:

  • Ito M  et al. Prostanoid-dependent spontaneous pain and PAR2-dependent mechanical allodynia following oral mucosal trauma: involvement of TRPV1, TRPA1 and TRPV4. Mol Pain 13:1744806917704138 (2017). Read more (PubMed: 28381109) »
  • Vieira ML  et al. Modulation of Hemostatic and Inflammatory Responses by Leptospira Spp. PLoS Negl Trop Dis 10:e0004713 (2016). Read more (PubMed: 27167223) »
See all 2 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Detects Rhesus Bradykinin

Good Average 3/5 (Ease of Use)
Abreviews
Objective:
We used this kit to test bradykinin concentrations in serum and EDTA plasma samples from human and rhesus macaque. No samples were Ethanol treated. We also performed a spike recovery on diluent and both rhesus serum and plasma

Ease of Use:
We were confused on some issues regarding the controls and methods of the procedures. There have been several versions of the SOP in recent times, and we found that Version 3, which came with the kit, had already been supplanted by Version 4 and 5 within months. The directions and recommendations varied. The EDTA plasma preparation recommendations are confusing: "Mix blood in a ratio of 1:4 with ice cold ethanol." Should this be 1 part with 4 parts, or 1 part of 4 parts? "Dry down sample..." What? How? How can it be prevented from degrading during this "dry-down"? The different treatments for the control wells add points of common error, and the explanation on how to interpret these controls is not readily understood, particularly to those without experience with competitive ELISAs.

Interpretation: This kit does detect Rhesus bradykinin

We were able to determine that decreasing OD (competitive ELISA) correlated to increasing concentrations of Bradykinin in human and rhesus samples in a similar fashion.

Results: EDTA plasma:
For n=2 rhesus animals, extrapolated values were the same for dilutions of 1:16, 1:64, and 1:256 n=1 rhesus animal extrapolated values varied at all dilutions. In the corresponding human sample, results for 1:16 and 1:64 corresponded.

Results: Serum:
There seemed to be no correlation among the tested dilutions for extrapolated values in human or rhesus. We believe this relates to dilution-dependent ex vivo generation during the assay.

Spike Recover:
There was virtually no relationship between the amount spiked and the amount measured in the rhesus serum or plasma (notice the logarithmic scale for that test). We did not repeat this assay with human serum or plasma.

Values in these graphs are average +/- 1 standard deviation


ab136936: ABTRIAL-TVB33 value: $573

Abcam user community

Verified customer

Submitted Aug 25 2015

Answer

It is preferable to perform the extraction/handling as it is laid out in the kit protocol as this will help to remove interference as well as other factors that can contribute to the instability of the sample.

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Question
Answer



A pesar de que hayáis optimizado las diluciones para vuestras muestras, las diluciones que habéis usado están por debajo del mínimo recomendado para evitar el efecto matriz. Os sugiero una dilución mínima de 1/16 para plasma y orina.
De los resultados enviados observamos un alto coeficiente de variación entre duplicados. Esto suele deberse a problemas ocurridos durante el pipeteo o a algo que se haya añadido al tampón de dilución que pueda estar interfiriendo en las medidas.
La curva de estándares es prácticamente plana, pero lo más preocupante es que varios de los puntos de la curva superan en absorbancia al valor de B0, lo cual es imposible en la práctica. B0 es el punto en el que el 100% del anticuerpo esta unido al estándar, y por tanto da el valor más alto de absorbancia en muestras y estándares, ya que no hay Bradykinina para competir por el anticuerpo.



El hecho de que este valor sea más bajo que otros puntos de la recta implica que algo ha ido mal durante el ensayo o la configuración de la placa durante el mismo.

¿Estáis usando algún otro diluyente aparte del Assay Buffer del kit para diluir el estándar?
Una vez preparada los estándares para la curva, ¿se tardaron más de 30 minutos en tomar las medidas de los mismos? En caso contrario el estándar pudo haberse degradado.
¿Se siguió el esquema de concentraciones de estándares indicado en el protocolo o se prepararon diluciones diferentes?
¿Estaban todos los reactivos a temperatura ambiente antes de usarlos, habiéndolos sacado durante al menos 30 minutos antes de comenzar el ensayo? En caso contrario la eficiencia de unión del anticuerpo puede disminuir.
¿Se eliminó completamente el buffer de lavado de los pocillos antes de añadir el reactivo? Es importante asegurarse que no quedan restos de buffer en los pocillos, pero teniendo cuidado de no secar la placa (si se invierte la placa y golpea para eliminar el exceso de buffer de lavado, debe hacerse rápido y con un par de golpecitos simplemente).



Os animo a repetir la curva de estándares, si aun es posible, y probar con varias diluciones de una muestra representativa para comprobar si los valores siguen sin ser los esperados. Aunque pueda parecer evidente también os recomiendo comprobar que la configuración de la placa es la adecuada y que el lector está configurado de forma apropiada para leer a la longitud de onda indicada.

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