Mouse RBP4 Quantikine ELISA Kit

(2 citations)   
  • 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), Cell Lysates (50 uL), Serum (10 uL), EDTA Plasma (10 uL), Heparin Plasma (10 uL), Urine (25 uL)
  • Sensitivity
    0.043 ng/mL
  • Assay Range
    0.6 - 20 ng/mL (Cell Culture Supernates, Cell Lysates, Serum, EDTA Plasma, Heparin Plasma, Urine)
  • Specificity
    Natural and recombinant mouse RBP4
  • Cross-reactivity
    < 0.5% cross-reactivity observed with available related molecules.< 50% cross-species reactivity observed with species tested.
  • Interference
    No significant interference observed with available related molecules.
Product Summary
The Quantikine Mouse RBP4 immunoassay is a 4.5 hour solid-phase ELISA designed to measure RBP4 in cell culture supernates, cell lysates, mouse serum, plasma, and urine. It contains NS0-expressed recombinant mouse RBP4 and antibodies raised against the recombinant factor. This immunoassay has been shown to accurately quantitate recombinant mouse RBP4. Results obtained using natural mouse RBP4 showed dose response curves that were parallel to the standard curve obtained using the Quantikine mouse kit standards. These results indicate that this kit can be used to determine relative mass values for natural mouse RBP4.

Precision
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision.
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in assays to assess inter-assay precision.

Intra-Assay Precision Inter-Assay Precision
Sample123123
n202020202020
Mean0.8962.239.440.9022.228.52
Standard Deviation0.0360.120.510.0890.150.64
CV%45.15.49.86.77.5

Cell Culture Supernates, Cell Lysates, Serum, EDTA Plasma, Heparin Plasma, Urine
Intra-Assay Precision Inter-Assay Precision
Sample123123
n202020202020
Mean0.8962.239.440.9022.228.52
Standard Deviation0.0360.120.510.0890.150.64
CV%45.45.49.96.87.5

Recovery

The recovery of mouse RBP4 spiked to three levels throughout the range of the assay was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Supernates (n=4) 98 90-112
Urine (n=4) 91 80-107
Linearity
To assess the linearity of the assay, samples containing and/or spiked with high concentrations of mouse RBP4 in each matrix were diluted with Calibrator Diluent and assayed.
Mouse RBP4 Quantikine ELISA Kit
Preparation and Storage
  • Storage
    Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: RBP4/Retinol-Binding Protein 4
Retinol-binding protein 4 (RBP4) is a Lipocalin superfamily molecule that transports vitamin A (retinol) and retinaldehyde in the serum. Retinaldehyde is the critical chromophore in the rhodopsin photoreceptor, while both it and retinoic acid regulate a multitude of cellular differentiation and proliferation effects through the intracellular receptors RAR and RXR. RBP4 is secreted primarily by hepatocytes and adipocytes into the blood where the RBP4-retinol complex interacts with transthyretin (TTR). Formation of the complex with TTR increases the serum half-life of RBP4 by preventing RBP4 filtration through the kidney. The C-terminally processed forms of RBP4, which do not bind TTR, are normally excreted into the urine but accumulate in the serum during renal failure. RBP4 promotes hyperglycemia, and its upregulation in visceral and liver adipocytes leads to elevated serum levels.
    • Entrez Gene IDs
      5950 (Human); 19662 (Mouse); 25703 (Rat);
    • Alternate Names
      interstitial; Plasma retinol-binding protein; RBP4; retinol binding protein 4, plasma; RetinolBinding Protein 4; retinol-binding protein 4, plasma;
    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.

    3. 50 µL Assay Diluent
    4.   Add 50 µL of Assay Diluent to each well.

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

    8. 100 µL Conjugate
    9.   Add 100 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
    10.   Aspirate and wash 5 times.

    11. 100 µL Substrate Solution
    12.   Add 100 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.

    13. 100 µL Stop Solution
    14.   Add 100 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
    Citations:

    R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

    2 Citations: Showing 1 - 2
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    Species
    Sample Type
    1. Transgenic Mice Over-Expressing RBP4 Have RBP4-Dependent and Light-Independent Retinal Degeneration
      Authors: M Du, E Phelps, MJ Balangue, A Dockins, G Moiseyev, Y Shin, S Kane, L Otalora, JX Ma, R Farjo, KM Farjo
      Invest. Ophthalmol. Vis. Sci., 2017;58(10):4095-4103.
      Species: Mouse
      Sample Type: Serum
    2. Serum retinol-binding protein-induced endothelial inflammation is mediated through the activation of toll-like receptor 4
      Authors: M Du, A Martin, F Hays, J Johnson, RA Farjo, KM Farjo
      Mol. Vis., 2017;23(0):185-197.
      Species: Mouse
      Sample Type: Serum
    Supplemental ELISA Products
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    Quantikine ELISA Wash Buffer 1

    ELISA WA126 6
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    Cell Lysis Buffer 2

    ELISA 895347 2
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