Human FGF23 ELISA Kit (HUES02207)

SKU:
HUES02207
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
Q9GZV9
Sensitivity:
9.38pg/mL
Range:
15.63-1000pg/mL
ELISA Type:
Sandwich
Synonyms:
FGF-23, ADHR, FGFN, HYPF, HPDR2, PHPTC
Reactivity:
Human
Tested Sample Types:
Serum, plasma and other biological fluids
€299

Description

ELISA Kit Technical ManualMSDS

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.

Assay type Sandwich
Format 96T
Assay time 4.5h
Reactivity Human
Detection Method Colormetric
Detection Range 15.63-1000 pg/mL
Sensitivity 9.38 pg/mL
Sample Volume Required Per Well 100uL
Sample Type Serum, plasma and other biological fluids

Specificity

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.

Concentration
(pg/mL)
O.DAverageCorrected
1000 2.415
2.429
2.422 2.341
500 1.563
1.621
1.592 1.511
250 0.918
0.892
0.905 0.824
125 0.434
0.454
0.444 0.363
62.5 0.276
0.258
0.267 0.186
31.25 0.183
0.177
0.18 0.099
15.63 0.123
0.141
0.132 0.051
0 0.071
0.091
0.081 --

Precision

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 PrecisionInter-assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 20 20 20
Mean (pg/mL) 50.86 87.87 437.81 48.92 79.96 443.72
Standard deviation 3.31 4.66 19.96 2.75 3.49 20.85
C V (%) 6.51 5.30 4.56 5.62 4.36 4.70

Recovery

The recovery of Human FGF23 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample TypeRange (%)Average Recovery (%)
Serum (n=5) 94-110 101
EDTA plasma (n=5) 96-107 102
Cell culture media (n=5) 89-104 95

Linearity

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)
1:2 Range (%) 94-108 96-111 89-103
Average (%) 101 102 96
1:4 Range (%) 92-105 81-94 86-98
Average (%) 99 88 92
1:8 Range (%) 91-104 85-101 87-101
Average (%) 98 92 93
1:16 Range (%) 93-107 86-96 81-96
Average (%) 99 91 88

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.

ItemSpecificationsStorage
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]
UniProt Code: Q9GZV9
NCBI GenInfo Identifier: 13626688
NCBI Gene ID: 8074
NCBI Accession: Q9GZV9.1
UniProt Secondary Accession: Q9GZV9,Q4V758,
UniProt Related Accession: Q9GZV9
Molecular Weight:
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