Human FGF23 ELISA Kit
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human FGF23 . Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human FGF23 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human FGF23, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human FGF23. The concentration of Human FGF23 in samples can be calculated by comparing the OD of the samples to the standard curve.
|Detection Range||15.63-1000 pg/mL|
|Sample Volume Required Per Well||100uL|
|Sample Type||Serum, plasma and other biological fluids|
This kit recognizes Human FGF23 in samples. No significant cross-reactivity or interference between Human FGF23 and analogues was observed.
FGF23 ELISA Kit Typical Data
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human FGF23 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human FGF23 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||6.51||5.30||4.56||5.62||4.36||4.70|
The recovery of Human FGF23 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||96-107||102|
|Cell culture media (n=5)||89-104||95|
Samples were spiked with high concentrations of Human FGF23 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
FGF23 ELISA Kit Components & Storage
An unopened kit can be stored at 4'C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
|Micro ELISA Plate(Dismountable)||8 wells X12 strips||-20'C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100X)||1 vial, 120 uL|
|Concentrated HRP Conjugate (100X)||1 vial, 120 uL||-20'C(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4'C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25X)||1 vial, 30 mL|
|Substrate Reagent||1 vial, 10 mL||4'C(shading light)|
|Stop Solution||1 vial, 10 mL||4'C|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
Human FGF23 ELISA Kit (HUES02207) Assay procedure
- 1. Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- 2. Aliquot 100µl of standard solutions into the standard wells.
- 3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- 4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- 5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- 6. Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix.Incubate for 1 hour at 37°C.
- 7. Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- 8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- 9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- 10. Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- 11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- 12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
Human FGF23 ELISA Kit (HUES02207) Protein Information
|UniProt Protein Function:||FGF23: Regulator of phosphate homeostasis. Inhibits renal tubular phosphate transport by reducing SLC34A1 levels. Upregulates EGR1 expression in the presence of KL. Acts directly on the parathyroid to decrease PTH secretion. Regulator of vitamin-D metabolism. Negatively regulates osteoblast differentiation and matrix mineralization. Defects in FGF23 are the cause of autosomal dominant hypophosphataemic rickets (ADHR). ADHR is characterized by low serum phosphorus concentrations, rickets, osteomalacia, leg deformities, short stature, bone pain and dental abscesses. Defects in FGF23 are a cause of hyperphosphatemic familial tumoral calcinosis (HFTC). HFTC is a severe autosomal recessive metabolic disorder that manifests with hyperphosphatemia and massive calcium deposits in the skin and subcutaneous tissues. Belongs to the heparin-binding growth factors family.|
|UniProt Protein Details:|| |
Protein type:Secreted, signal peptide; Secreted; Cytokine
Chromosomal Location of Human Ortholog: 12p13.3
Cellular Component: extracellular space; extracellular region
Molecular Function:growth factor activity; type 1 fibroblast growth factor receptor binding
Biological Process: epidermal growth factor receptor signaling pathway; negative regulation of bone mineralization; phosphoinositide-mediated signaling; fibroblast growth factor receptor signaling pathway; cellular phosphate ion homeostasis; nerve growth factor receptor signaling pathway; positive regulation of transcription, DNA-dependent; negative regulation of hormone secretion; insulin receptor signaling pathway; innate immune response; negative regulation of osteoblast differentiation; phosphate ion homeostasis; cell differentiation; phosphate metabolic process; vitamin D catabolic process
Disease: Hypophosphatemic Rickets, Autosomal Dominant
|NCBI Summary:||This gene encodes a member of the fibroblast growth factor family of proteins, which possess broad mitogenic and cell survival activities and are involved in a variety of biological processes. The product of this gene regulates phosphate homeostasis and transport in the kidney. The full-length, functional protein may be deactivated via cleavage into N-terminal and C-terminal chains. Mutation of this cleavage site causes autosomal dominant hypophosphatemic rickets (ADHR). Mutations in this gene are also associated with hyperphosphatemic familial tumoral calcinosis (HFTC). [provided by RefSeq, Feb 2013]|
|NCBI GenInfo Identifier:||13626688|
|NCBI Gene ID:||8074|
|UniProt Secondary Accession:||Q9GZV9,Q4V758,|
|UniProt Related Accession:||Q9GZV9|
|NCBI Full Name:||Fibroblast growth factor 23|
|NCBI Synonym Full Names:||fibroblast growth factor 23|
|NCBI Official Symbol:||FGF23|
|NCBI Official Synonym Symbols:||ADHR; FGFN; HYPF; HPDR2; PHPTC|
|NCBI Protein Information:||fibroblast growth factor 23; phosphatonin; tumor-derived hypophosphatemia inducing factor|
|UniProt Protein Name:||Fibroblast growth factor 23|
|UniProt Synonym Protein Names:||Phosphatonin; Tumor-derived hypophosphatemia-inducing factorCleaved into the following 2 chains:Fibroblast growth factor 23 N-terminal peptide; Fibroblast growth factor 23 C-terminal peptide|
|Protein Family:||Fibroblast growth factor|
|UniProt Gene Name:||FGF23|
|UniProt Entry Name:||FGF23_HUMAN|