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ZNF638 Colorimetric Cell-Based ELISA

SKU:
CBCAB01100
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

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ZNF638 Colorimetric Cell-Based ELISA

The ZNF638 Colorimetric Cell-Based ELISA Kit is a cutting-edge assay designed for the precise measurement of Zinc Finger Protein 638 (ZNF638) levels in cell lysates and culture supernatants. This innovative kit offers exceptional sensitivity and specificity, ensuring accurate and reproducible results for a variety of research applications.ZNF638 is a key regulatory protein that plays a crucial role in gene expression, cell growth, and DNA repair mechanisms. Dysregulation of ZNF638 has been implicated in various diseases, including cancer, autoimmune disorders, and developmental abnormalities, making it a vital target for therapeutic interventions and biomarker studies.

With its user-friendly protocol and high-performance capabilities, the ZNF638 Colorimetric Cell-Based ELISA Kit is an indispensable tool for researchers seeking to unravel the intricate mechanisms underlying ZNF638 function and its implications in disease pathogenesis. Unlock the potential of your research with this advanced ELISA kit from Assay Genie.

Product Name:ZNF638 Colorimetric Cell-Based ELISA
Product Code:CBCAB01100
ELISA Type:Cell-Based
Target:ZNF638
Reactivity:Human
Dynamic Range:> 5000 Cells
Detection Method:Colorimetric 450 nmStorage/Stability:4°C/6 Months
Format:96-Well Microplate

The ZNF638 Colorimetric Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can detect ZNF638 protein expression profile in cells. The kit can be used for measuring the relative amounts of ZNF638 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie siRNA or chemicals), or activators have on ZNF638.

Qualitative determination of ZNF638 concentration is achieved by an indirect ELISA format. In essence, ZNF638 is captured by ZNF638-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: 27332, UniProt ID: Q14966, OMIM: 0, Unigene: Hs.434401
Gene Symbol:ZNF638
Sub Type:None
UniProt Protein Function:ZNF638: Early regulator of adipogenesis that works as a transcription cofactor of CEBPs, controlling the expression of PPARG and probably of other proadipogenic genes, such as SREBF1. Binds to cytidine clusters in double-stranded DNA. 5 isoforms of the human protein are produced by alternative splicing.
UniProt Protein Details:

Protein type:DNA-binding; RNA-binding

Chromosomal Location of Human Ortholog: 2p13.1

Cellular Component: nucleoplasm; intracellular membrane-bound organelle; cytoplasm; nuclear speck

Molecular Function:zinc ion binding; RNA binding; double-stranded DNA binding; nucleotide binding

Biological Process: transcription, DNA-dependent; regulation of transcription, DNA-dependent; RNA splicing

NCBI Summary:The protein encoded by this gene is a nucleoplasmic protein. It binds cytidine-rich sequences in double-stranded DNA. This protein has three types of domains: MH1, MH2 (repeated three times) and MH3. It is associated with packaging, transferring, or processing transcripts. Multiple alternatively spliced transcript variants have been found for this gene, but the biological validity of some variants has not been determined. [provided by RefSeq, Jul 2008]
UniProt Code:Q14966
NCBI GenInfo Identifier:62526045
NCBI Gene ID:27332
NCBI Accession:NP_001014972.1
UniProt Related Accession:Q14966
Molecular Weight:
NCBI Full Name:zinc finger protein 638 isoform 1
NCBI Synonym Full Names:zinc finger protein 638
NCBI Official Symbol:ZNF638  
NCBI Official Synonym Symbols:ZFML; NP220; Zfp638  
NCBI Protein Information:zinc finger protein 638
UniProt Protein Name:Zinc finger protein 638
UniProt Synonym Protein Names:Cutaneous T-cell lymphoma-associated antigen se33-1; CTCL-associated antigen se33-1; Nuclear protein 220; Zinc finger matrin-like protein
UniProt Gene Name:ZNF638  
UniProt Entry Name:ZN638_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-ZNF638 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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