Human GATA4 (GATA Binding Protein 4) 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 GATA4 . 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 GATA4 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 GATA4, 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 GATA4. The concentration of Human GATA4 in samples can be calculated by comparing the OD of the samples to the standard curve.
|Detection Range||1.56-100 ng/mL|
|Sample Volume Required Per Well||100uL|
|Sample Type||Serum, plasma and other biological fluids|
This kit recognizes Human GATA4 in samples. No significant cross-reactivity or interference between Human GATA4 and analogues was observed.
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 GATA4 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human GATA4 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||6.76||5.00||3.97||5.76||5.77||4.04|
The recovery of Human GATA4 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)||91-103||98|
|Cell culture media (n=5)||95-106||101|
Samples were spiked with high concentrations of Human GATA4 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)|
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 GATA4 (GATA Binding Protein 4) ELISA Kit (HUES02010) 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 GATA4 (GATA Binding Protein 4) ELISA Kit (HUES02010) Protein Information
|UniProt Protein Function:||GATA4: a conserved and ubiquitous member of the GATA family of zinc-finger transcription factors. Members of this family recognize the GATA motif which is present in the promoters of many genes. This protein is thought to regulate genes involved in embryogenesis and in myocardial differentiation and function. May regulate a set of cardiac-specific genes and play a crucial role in cardiogenesis. Defects are a cause of atrial septal defect 2 (ASD2). ASD2 is an autosomal dominant condition with atrial septal defect and other congenital heart disease but no conduction defects or noncardiac abnormalities.|
|UniProt Protein Details:|
Protein type:Motility/polarity/chemotaxis; Transcription factor
Chromosomal Location of Human Ortholog: 8p23.1-p22
Cellular Component: nucleoplasm; nucleus
Molecular Function:chromatin binding; DNA binding; protein binding; sequence-specific DNA binding; transcription activator binding; transcription coactivator activity; transcription factor activity; transcription factor binding
Biological Process: anatomical structure formation; blood coagulation; cell development; cell fate commitment; cell-cell signaling; embryonic foregut morphogenesis; embryonic heart tube anterior/posterior pattern formation; endoderm development; heart looping; male gonad development; positive regulation of angiogenesis; positive regulation of cardioblast differentiation; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; regulation of transcription, DNA-dependent; response to drug; transcription from RNA polymerase II promoter
Disease: Atrial Septal Defect 2; Atrioventricular Septal Defect 4; Testicular Anomalies With Or Without Congenital Heart Disease; Tetralogy Of Fallot; Ventricular Septal Defect 1
|NCBI Summary:||This gene encodes a member of the GATA family of zinc-finger transcription factors. Members of this family recognize the GATA motif which is present in the promoters of many genes. This protein is thought to regulate genes involved in embryogenesis and in myocardial differentiation and function, and is necessary for normal testicular development. Mutations in this gene have been associated with cardiac septal defects. Additionally, alterations in gene expression have been associated with several cancer types. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Apr 2015]|
|NCBI GenInfo Identifier:||215274105|
|NCBI Gene ID:||2626|
|UniProt Secondary Accession:||P43694,Q3MJ45, Q5IFM8, B7ZKX0, B7ZKZ4,|
|UniProt Related Accession:||P43694|
|Molecular Weight:||44,665 Da|
|NCBI Full Name:||Transcription factor GATA-4|
|NCBI Synonym Full Names:||GATA binding protein 4|
|NCBI Official Symbol:||GATA4|
|NCBI Official Synonym Symbols:||TOF; ASD2; VSD1; TACHD|
|NCBI Protein Information:||transcription factor GATA-4|
|UniProt Protein Name:||Transcription factor GATA-4|
|UniProt Synonym Protein Names:||GATA-binding factor 4|
|Protein Family:||GATA transcription factor|
|UniProt Gene Name:||GATA4|
|UniProt Entry Name:||GATA4_HUMAN|