Human SOX3 (Sex Determining Region Y Box Protein 3) ELISA Kit (HUES03045)

SKU:
HUES03045
€539

Description

ELISA Kit Technical ManualMSDS

Human SOX3 (Sex Determining Region Y Box Protein 3) 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 SOX3 . 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 SOX3 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 SOX3, 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 SOX3. The concentration of Human SOX3 in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityHuman
Detection MethodColormetric
Detection Range78.13-5000 pg/mL
Sensitivity46.88 pg/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human SOX3 in samples. No significant cross-reactivity or interference between Human SOX3 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.

Concentration
(pg/mL)
O.D Average Corrected
50002.442
2.5
2.4712.381
25001.741
1.779
1.761.67
12501.044
1.006
1.0250.935
6250.495
0.519
0.5070.417
312.50.318
0.316
0.3170.227
156.250.201
0.195
0.1980.108
78.130.141
0.149
0.1450.055
00.083
0.097
0.09--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human SOX3 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human SOX3 were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)272.43462.862102.73291.37478.822221.44
Standard deviation15.5023.2480.7418.9725.3398.63
C V (%)5.695.023.846.515.294.44

Recovery

The recovery of Human SOX3 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)89-10194
EDTA plasma (n=5)89-10496
Cell culture media (n=5)87-10394

Linearity

Samples were spiked with high concentrations of Human SOX3 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 (%)94-10889-10398-115
Average (%)10196105
1:4Range (%)87-9984-9989-103
Average (%)939094
1:8Range (%)88-10082-9284-98
Average (%)948691
1:16Range (%)94-10682-9581-94
Average (%)1008787

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 SOX3 (Sex Determining Region Y Box Protein 3) ELISA Kit (HUES03045) 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 SOX3 (Sex Determining Region Y Box Protein 3) ELISA Kit (HUES03045) Protein Information

UniProt Protein Function:SOX3: Transcription factor required during the formation of the hypothalamo-pituitary axis. May function as a switch in neuronal development. Keeps neural cells undifferentiated by counteracting the activity of proneural proteins and suppresses neuronal differentiation. Required also within the pharyngeal epithelia for craniofacial morphogenesis. Controls a genetic switch in male development. Is necessary for initiating male sex determination by directing the development of supporting cell precursors (pre-Sertoli cells) as Sertoli rather than granulosa cells. Defects in SOX3 are a cause of panhypopituitarism X- linked (PHPX). Affected individuals have absent infundibulum, anterior pituitary hypoplasia, and ectopic posterior pituitary. Defects in SOX3 are the cause of mental retardation X- linked with isolated growth hormone deficiency (MRXGH). Defects in SOX3 are the cause of 46,XX sex reversal type 3 (SRXX3). A condition in which male gonads develop in a genetic female (female to male sex reversal). Copy number variations (CNV) encompassing or in close proximity to SOX3 are responsible for XX male reversal. These variations include two duplications of approximately 123 kb and 85 kb, the former of which spans the entire SOX3 gene; a 343 kb deletion immediately upstream of SOX3 that is probably responsible of altered regulation (and not increased dosage) of SOX3; a large (approximately 6 Mb) duplication that encompasses SOX3 and at least 18 additional distally located genes. Its proximal breakpoint falls within the SOX3 regulatory region. This large rearrangement has been found in a patient with XX male reversal and a complex phenotype that also includes a scrotal hypoplasia, microcephaly, developmental delay, and growth retardation.
UniProt Protein Details:

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

Chromosomal Location of Human Ortholog: Xq27.1

Cellular Component: nucleoplasm

Molecular Function:DNA binding

Biological Process: central nervous system development; hypothalamus development; negative regulation of neuron differentiation; pituitary gland development; sensory organ development; sex determination

Disease: Mental Retardation, X-linked, With Panhypopituitarism; Panhypopituitarism, X-linked

NCBI Summary:This gene encodes a member of the SOX (SRY-related HMG-box) family of transcription factors involved in the regulation of embryonic development and in the determination of the cell fate. The encoded protein may act as a transcriptional regulator after forming a protein complex with other proteins. Mutations in this gene have been associated with X-linked mental retardation with growth hormone deficiency. [provided by RefSeq, Jul 2008]
UniProt Code:P41225
NCBI GenInfo Identifier:48429228
NCBI Gene ID:6658
NCBI Accession:P41225.2
UniProt Secondary Accession:P41225,P35714, Q5JWI3, Q9NP49,
UniProt Related Accession:P41225
Molecular Weight:45,210 Da
NCBI Full Name:Transcription factor SOX-3
NCBI Synonym Full Names:SRY-box 3
NCBI Official Symbol:SOX3
NCBI Official Synonym Symbols:PHP; GHDX; MRGH; PHPX; SOXB
NCBI Protein Information:transcription factor SOX-3
UniProt Protein Name:Transcription factor SOX-3
Protein Family:Transcription factor
UniProt Gene Name:SOX3
UniProt Entry Name:SOX3_HUMAN
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Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
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