Human FOXO1 (Forkhead Box Protein O1) ELISA Kit (HUES02193)



ELISA Kit Technical ManualMSDS

Human FOXO1 (Forkhead Box Protein O1) 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 FOXO1 . 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 FOXO1 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 FOXO1, 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 FOXO1. The concentration of Human FOXO1 in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Assay time4.5h
Detection MethodColormetric
Detection Range0.16-10 ng/mL
Sensitivity0.10 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids


This kit recognizes Human FOXO1 in samples. No significant cross-reactivity or interference between Human FOXO1 and analogues was observed.

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.

O.D Average Corrected


Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human FOXO1 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human FOXO1 were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Mean (ng/mL)0.551.054.230.561.013.96
Standard deviation0.
C V (%)5.454.765.205.364.954.04


The recovery of Human FOXO1 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)88-10295
EDTA plasma (n=5)87-9993
Cell culture media (n=5)88-10194


Samples were spiked with high concentrations of Human FOXO1 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)
1:2Range (%)91-10685-10091-105
Average (%)989198
1:4Range (%)92-10382-9785-98
Average (%)978991
1:8Range (%)88-10385-9987-97
Average (%)959192
1:16Range (%)91-10684-9889-101
Average (%)978995

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.

Item Specifications Storage
Micro ELISA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 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 Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Human FOXO1 (Forkhead Box Protein O1) ELISA Kit (HUES02193) 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 100 µL of standard solutions into the standard wells.
    3. Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100 µL 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 100 µL 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 350 µL 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 100 µL 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 90 µL 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 FOXO1 (Forkhead Box Protein O1) ELISA Kit (HUES02193) Protein Information

UniProt Protein Function:FOXO1A: a transcription factor of the forkhead family. A central regulator of metabolism in several cell types. May play an important role in myogenic growth and differentiation, and in the regulation of lipolysis in adipocytes by controlling the expression of adipose triglyceride lipase (ATGL), the rate-limiting lipolytic enzyme. Translocation of this gene with PAX3 has been associated with alveolar rhabdomyosarcoma. Contains 1 fork-head domain.
UniProt Protein Details:

Protein type:Nuclear receptor co-regulator; Oncoprotein; DNA-binding; Transcription factor

Chromosomal Location of Human Ortholog: 13q14.1

Cellular Component: cytoplasm; cytosol; mitochondrion; nucleoplasm; nucleus

Molecular Function:beta-catenin binding; chromatin binding; protein binding; protein phosphatase 2A binding; sequence-specific DNA binding; ubiquitin protein ligase binding

Biological Process: anatomical structure morphogenesis; apoptosis; autophagy; blood vessel development; cell differentiation; cell glucose homeostasis; cellular response to insulin stimulus; cellular response to starvation; endocrine pancreas development; epidermal growth factor receptor signaling pathway; fat cell differentiation; fibroblast growth factor receptor signaling pathway; innate immune response; insulin receptor signaling pathway; negative regulation of apoptosis; negative regulation of fat cell differentiation; negative regulation of stress-activated MAPK cascade; negative regulation of transcription from RNA polymerase II promoter; negative regulation of transcription, DNA-dependent; nerve growth factor receptor signaling pathway; phosphoinositide-mediated signaling; positive regulation of apoptosis; positive regulation of autophagy; positive regulation of gluconeogenesis; positive regulation of protein catabolic process; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; protein amino acid acetylation; response to DNA damage stimulus; thermoregulation; transcription, DNA-dependent

Disease: Rhabdomyosarcoma 2

NCBI Summary:This gene belongs to the forkhead family of transcription factors which are characterized by a distinct forkhead domain. The specific function of this gene has not yet been determined; however, it may play a role in myogenic growth and differentiation. Translocation of this gene with PAX3 has been associated with alveolar rhabdomyosarcoma. [provided by RefSeq, Jul 2008]
UniProt Code:Q12778
NCBI GenInfo Identifier:116241368
NCBI Gene ID:2308
NCBI Accession:Q12778.2
UniProt Secondary Accession:Q12778,O43523, Q5VYC7, Q6NSK6,
UniProt Related Accession:Q12778
Molecular Weight:
NCBI Full Name:Forkhead box protein O1
NCBI Synonym Full Names:forkhead box O1
NCBI Official Symbol:FOXO1
NCBI Official Synonym Symbols:FKH1; FKHR; FOXO1A
NCBI Protein Information:forkhead box protein O1
UniProt Protein Name:Forkhead box protein O1
UniProt Synonym Protein Names:Forkhead box protein O1A; Forkhead in rhabdomyosarcoma
Protein Family:Foxo1-corepressor
UniProt Gene Name:FOXO1
UniProt Entry Name:FOXO1_HUMAN
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