Human/Canine/Porcine Insulin Quantikine ELISA Kit

  
  • Assay Type
    Solid Phase Sandwich ELISA
  • Format
    96-well strip plate
  • Assay Length
    4.5 hours
  • Sample Type & Volume Required Per Well
    Cell Culture Supernates (50 uL), Serum (50 uL), EDTA Plasma (50 uL), Heparin Plasma (50 uL)
  • Sensitivity
    2.15 pmol/L
  • Assay Range
    15.6 - 500 pmol/L (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma)
  • Specificity
    Recombinant human Insulin and natural human, canine, and porcine Insulin.  This assay cross-reacts 0.25% with Human Proinsulin.
  • Cross-reactivity
    < 0.5% cross-reactivity observed with available related molecules.Cross-species reactivity observed with 1 or more species tested.
  • Interference
    No significant interference observed with available related molecules.
Control Available
QC107, Quantikine Immunoassay Control Set 819 for Insulin - Please Inquire
Product Summary
The Quantikine Human/Canine/Porcine Insulin Immunoassay is a 4.5 hour solid-phase ELISA designed to measure Insulin in cell culture supernates, serum, and plasma. It contains S. cerevisiae-expressed recombinant human Insulin and has been shown to accurately quantitate the recombinant factor. Results obtained using natural Insulin showed linear curves that were parallel to the standard curves obtained using the Quantikine kit standards. These results indicate that this kit can be used to determine relative mass values for naturally occurring Insulin.

Precision
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in twenty separate assays to assess inter-assay precision. Assays were performed by at least three technicians using two lots of components.
Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma
Intra-Assay Precision Inter-Assay Precision
Sample123123
n202020202020
Mean66.815829962.2148288
Standard Deviation2.585.8911.94.610.221.5
CV%3.93.747.46.97.5

Recovery

The recovery of Insulin spiked to levels throughout the range of the assay in various matrices was evaluated

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 95 85-108
EDTA Plasma (n=4) 95 86-102
Heparin Plasma (n=4) 92 85-105
Serum (n=4) 96 87-103
Linearity
To assess the linearity of the assay, samples containing and/or spiked with high concentrations of Insulin were diluted with Calibrator Diluent to produce samples with values within the dynamic range of the assay.
Human/Canine/Porcine Insulin Quantikine ELISA Kit
Preparation and Storage
  • Storage
    Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: Insulin
Insulin and the IGFs are members of the insulin family of molecules. IGF-I and II are structurally homologous to proinsulin, and can be thought of as taking the shape of an exaggerated, inverted letter G. Intrachain disulfide bonds link sections of the G. The single chain insulin propeptide consists of a 30 amino acid B chain (aa 2554), a C-peptide (aa 5589), and a 21 aa A chain (aa 90110). Removal of the C-peptide by proteolysis enables the formation of mature Insulin, a disulfidelinked heterodimer of the A and B chains. Circulating C-peptide levels are elevated in hyperinsulinism, obesity, and type II diabetes.
    • Entrez Gene IDs
      3630 (Human); 16333 (Mouse); 397415 (Porcine); 280829 (Bovine); 483665 (Canine);
    • Alternate Names
      IDDM2; ILPR; INS; Insulin; IRDN; MODY10; proinsulin;
    Related Research Areas
    Assay Procedure
    Refer to the product for complete assay procedure.

    Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
    1.   Prepare all reagents, standard dilutions, and samples as directed in the product insert.
    2.   Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
      1.   Aspirate each well and wash, repeating once more for a total of 2 washes.

      2. 150 µL Assay Diluent
      3.   Add 150 µL of Assay Diluent to each well.

      4. 50 µL Standard, Control, or Sample
      5.   Add 50 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours on a horizontal orbital microplate shaker.
      6.   Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.

      7. 200 µL Conjugate
      8.   Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours on the shaker.
      9.   Aspirate and wash 4 times.

      10. 200 µL Substrate Solution
      11.   Add 200 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes on the benchtop. PROTECT FROM LIGHT.

      12. 50 µL Stop Solution
      13.   Add 50 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
    ELISA Controls
    Description Application Cat# Citations Images  

    Quantikine Immunoassay Control Set 819 for Insulin

    Ctrl QC107
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    Supplemental ELISA Products
    Description Application Cat# Citations Images  

    Quantikine ELISA Wash Buffer 1

    ELISA WA126 6
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