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ELF1 Colorimetric Cell-Based ELISA

SKU:
CBCAB01001
Product Type:
ELISA Kit
ELISA Type:
Cell Based
Research Area:
Epigenetics and Nuclear Signaling
Reactivity:
Human
Detection Method:
Colorimetric
€599
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Description

system_update_altDatasheet

ELF1 Colorimetric Cell-Based ELISA

The ELF1 Colorimetric Cell-Based ELISA Kit offered by AssayGenie is designed for the precise quantification of ELF1 (E74-like factor 1) protein levels in cell lysates and tissue homogenates. This kit provides high sensitivity and specificity, ensuring accurate and reproducible results for a variety of research applications.ELF1 is a transcription factor that plays a crucial role in regulating gene expression and cell growth. Dysregulation of ELF1 has been implicated in various diseases, including cancer and inflammatory disorders, making it a valuable biomarker for studying disease mechanisms and potential therapeutic targets.

With the ELF1 Colorimetric Cell-Based ELISA Kit, researchers can confidently measure ELF1 levels in their samples, providing valuable insights into cellular processes and disease pathways. Upgrade your research capabilities with this innovative kit from AssayGenie.

Product Name:ELF1 Colorimetric Cell-Based ELISA
Product Code:CBCAB01001
ELISA Type:Cell-Based
Target:ELF1
Reactivity:Human
Dynamic Range:> 5000 Cells
Detection Method:Colorimetric 450 nmStorage/Stability:4°C/6 Months
Format:96-Well Microplate

The ELF1 Colorimetric Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can detect ELF1 protein expression profile in cells. The kit can be used for measuring the relative amounts of ELF1 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie siRNA or chemicals), or activators have on ELF1.

Qualitative determination of ELF1 concentration is achieved by an indirect ELISA format. In essence, ELF1 is captured by ELF1-specific primary antibodies while the HRP-conjugated secondary antibodies bind the Fc region of the primary antibody. Through this binding, the HRP enzyme conjugated to the secondary antibody can catalyze a colorimetric reaction upon substrate addition. Due to the qualitative nature of the Cell-Based ELISA, multiple normalization methods are needed:

1. A monoclonal antibody specific for human GAPDH is included to serve as an internal positive control in normalizing the target absorbance values.
2. Following the colorimetric measurement of HRP activity via substrate addition, the Crystal Violet whole-cell staining method may be used to determine cell density. After staining, the results can be analysed by normalizing the absorbance values to cell amounts, by which the plating difference can be adjusted.
Database Information:Gene ID: 1997, UniProt ID: P32519, OMIM: 189973, Unigene: Hs.135646
Gene Symbol:ELF1
Sub Type:None
UniProt Protein Function:Elf-1: Transcription factor that activates the LYN and BLK promoters. Appears to be required for the T-cell-receptor-mediated trans activation of HIV-2 gene expression. Binds specifically to two purine-rich motifs in the HIV-2 enhancer. Belongs to the ETS family.
UniProt Protein Details:

Protein type:DNA-binding; Transcription factor

Chromosomal Location of Human Ortholog: 13q13

Cellular Component: nucleoplasm; nucleus

Molecular Function:protein binding; transcription factor activity

Biological Process: transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; negative regulation of T cell receptor signaling pathway; positive regulation of transcription from RNA polymerase II promoter; cell differentiation; regulation of cytokine production

NCBI Summary:This gene encodes an E26 transformation-specific related transcription factor. The encoded protein is primarily expressed in lymphoid cells and acts as both an enhancer and a repressor to regulate transcription of various genes. Alternative splicing results in multiple transcript variants. [provided by RefSeq, Feb 2009]
UniProt Code:P32519
NCBI GenInfo Identifier:212288097
NCBI Gene ID:1997
NCBI Accession:P32519.2
UniProt Secondary Accession:P32519,Q8N6F6, Q9UDE1, B4E2I5, E9PDQ9,
UniProt Related Accession:P32519
Molecular Weight:64,929 Da
NCBI Full Name:ETS-related transcription factor Elf-1
NCBI Synonym Full Names:E74-like factor 1 (ets domain transcription factor)
NCBI Official Symbol:ELF1  
NCBI Protein Information:ETS-related transcription factor Elf-1
UniProt Protein Name:ETS-related transcription factor Elf-1
UniProt Synonym Protein Names:E74-like factor 1
Protein Family:ETS-related transcription factor
UniProt Gene Name:ELF1  
UniProt Entry Name:ELF1_HUMAN
ComponentQuantity
96-Well Cell Culture Clear-Bottom Microplate2 plates
10X TBS24 mL
Quenching Buffer24 mL
Blocking Buffer50 mL
15X Wash Buffer50 mL
Primary Antibody Diluent12 mL
100x Anti-Phospho Target Antibody 60 µL
100x Anti-Target Antibody60 µL
Anti-GAPDH Antibody60 µL
HRP-Conjugated Anti-Rabbit IgG Antibody12 mL
HRP-Conjugated Anti-Mouse IgG Antibody12 mL
SDS Solution12 mL
Stop Solution24 mL
Ready-to-Use Substrate12 mL
Crystal Violet Solution12 mL
Adhesive Plate Seals2 seals

The following materials and/or equipment are NOT provided in this kit but are necessary to successfully conduct the experiment:

  • Microplate reader able to measure absorbance at 450 nm and/or 595 nm for Crystal Violet Cell Staining (Optional)
  • Micropipettes with capability of measuring volumes ranging from 1 µL to 1 ml
  • 37% formaldehyde (Sigma Cat# F-8775) or formaldehyde from other sources
  • Squirt bottle, manifold dispenser, multichannel pipette reservoir or automated microplate washer
  • Graph paper or computer software capable of generating or displaying logarithmic functions
  • Absorbent papers or vacuum aspirator
  • Test tubes or microfuge tubes capable of storing ≥1 ml
  • Poly-L-Lysine (Sigma Cat# P4832 for suspension cells)
  • Orbital shaker (optional)
  • Deionized or sterile water

*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

StepProcedure
1.Seed 200 µL of 20,000 adherent cells in culture medium in each well of a 96-well plate. The plates included in the kit are sterile and treated for cell culture. For suspension cells and loosely attached cells, coat the plates with 100 µL of 10 µg/ml Poly-L-Lysine (not included) to each well of a 96-well plate for 30 minutes at 37°C prior to adding cells.
2.Incubate the cells for overnight at 37°C, 5% CO2.
3.Treat the cells as desired.
4.Remove the cell culture medium and rinse with 200 µL of 1x TBS, twice.
5.Fix the cells by incubating with 100 µL of Fixing Solution for 20 minutes at room temperature. The 4% formaldehyde is used for adherent cells and 8% formaldehyde is used for suspension cells and loosely attached cells.
6.Remove the Fixing Solution and wash the plate 3 times with 200 µL 1x Wash Buffer for five minutes each time with gentle shaking on the orbital shaker. The plate can be stored at 4°C for a week.
7.Add 100 µL of Quenching Buffer and incubate for 20 minutes at room temperature.
8.Wash the plate 3 times with 1x Wash Buffer for 5 minutes each time.
9.Add 200 µL of Blocking Buffer and incubate for 1 hour at room temperature.
10. Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
11.Add 50 µL of 1x primary antibodies (Anti-ELF1 Antibody and/or Anti-GAPDH Antibody) to the corresponding wells, cover with Parafilm and incubate for 16 hours (overnight) at 4°C. If the target expression is known to be high, incubate for 2 hours at room temperature.
12.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
13.Add 50 µL of 1x secondary antibodies (HRP-Conjugated AntiRabbit IgG Antibody or HRP-Conjugated Anti-Mouse IgG Antibody) to corresponding wells and incubate for 1.5 hours at room temperature.
14.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
15.Add 50 µL of Ready-to-Use Substrate to each well and incubate for 30 minutes at room temperature in the dark.
16.Add 50 µL of Stop Solution to each well and read OD at 450 nm immediately using the microplate reader.

(Additional Crystal Violet staining may be performed if desired – details of this may be found in the kit technical manual.)

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