Antigen InformationAccession Number:P15976Gene ID:2623Gene Symbols:ERYF1GATA1GF1Protein Name & Synonyms:Erythroid transcription factor Eryf1 GATA-binding factor 1 (GATA-1, GF-1) NF-E1 DNA-binding proteinTarget Species:HumanAssay FormatSpecificity:The olionucleotide/antibody pair provided in this kit recognizes human GATA-1 in whole lysates and nuclear extracts.Number of Targets Detected:1Species Detected:HumanCompatible Sample Types:Cell LysatesNuclear ExtractsDesign Principle:Sandwich-basedMethod of Detection:ColorimetricQuantitative/Semi-Quantitative:Semi-QuantitativeSolid Support:96-well MicroplateProduct SpecificationsSize:1, 2, or 5 x 96-Well Strip Microplate KitIntroductionGATA transcription factors have a characteristic feature that they all bind to the DNA consensus sequence (A/T)GATA(A/G) of target gene promoters by two adjacent zinc fingers (Cys-X2-Cys-X17-Cys-X2-Cys). These transcription factors regulate differentiation, growth and survival of a wide range of cell types. The GATA family consists of six transcription factors, GATA1 to GATA6, between which the DNA-binding regions, two zinc finger domains are highly conserved. The C-terminal €˜€˜C-finger€™€™ and its adjacent basic region are necessary and sufficient for GATA to bind its cognate sequence, The N-terminal €˜€˜N-finger€™€™ can also bind DNA independently but has a preference for GATC core motifs. Both fingers participate in binding to the palindromic GATA motif ATCWGATA (W = A/T), resulting in markedly increased affinity.Product FeaturesSpecific transcription factor-DNA binding assayPerfect alternative to EMSAEasy to perform in an ELISA formatNon-radioactive assayHigh throughput (96-well plate format)Assay can be completed within 5 hoursApplication NotesKit Components96-well Strip Microplate pre-coated with DNA probesDNA Binding BufferPositive Control SampleSpecific Competitor DNA probeNon-specific Competitor DNA probeAssay ReagentDTTWash BufferPrimary AntibodyHRP-conjugated Secondary AntibodyAntibody Diluent BufferTMB One-Step Substrate ReagentStop SolutionOther Materials RequiredDistilled or deionized water100 ml and 1 liter graduated cylindersTubes to prepare sample dilutionsAbsorbent paperPrecision pipettes to deliver 2 µl to 1 ml volumesAdjustable 1-25 ml pipettes for reagent preparation< li="">Microplate reader capable of measuring absorbance at 450 nmProtocol OutlinePrepare all reagents and samples as instructed in the manual.Add 100 µl of sample or positive control to each well.Incubate 2 h at RT or O/N at 4 °C.Add 100 µl of prepared primary antibody to each well.Incubate 1 h at RT.Add 100 µl of prepared HRP-secondary antibody to each well.Incubate 1 h at RT.Add 100 µl of TMB One-Step Substrate Reagent to each well.Incubate 30 min at RT.Add 50 µl of Stop Solution to each well.Read at 450 nm immediately.Typical DataFigure 1Transcription factor activity assay of GATA-1 from nuclear extracts of K562 cells or HeLa cells. In K562 cells, activated GATA-1 is translocated into the nucleus where it binds with its corresponding DNA. A. Western-blot result of GATA-1 from cytoplasmic and nuclear fractions. B. Transcription factor activity assay of GATA-1 from nuclear fractions with the RayBio® GATA-1 Transcription Factor-Activity Assay Kit.Storage/StabilityUpon receipt, the positive control should be removed and stored at -20° or -80°C. The remainder of the kit can be stored for up to 6 months at 2-8°C from the date of shipment. Opened Microplate Wells or reagents may be stored for up to 1 month at 2° to 8°C. Return unused wells to the pouch containing desiccant pack, reseal along entire edge. Note: The kit can be used within one year if the whole kit is stored at -20°C upon receipt. Avoid repeated freeze-thaw cycles.No posts foundWrite A ReviewManualSDS