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CKI-gamma2 Colorimetric Cell-Based ELISA

SKU:
CBCAB01050
Product Type:
ELISA Kit
ELISA Type:
Cell Based
Research Area:
Metabolism
Reactivity:
Human
Mouse
Rat
Detection Method:
Colorimetric
€599
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Description

system_update_altDatasheetMSDS

CKI-gamma2 Colorimetric Cell-Based ELISA

The CKI Gamma2 Colorimetric Cell-Based ELISA Kit is specifically designed for the sensitive detection and quantification of CKI gamma2 protein levels in cell lysates. This kit offers high precision and accuracy, allowing for reliable and reproducible results for a variety of research applications.CKI gamma2 is a key regulator of cell cycle progression and cell division, playing a crucial role in maintaining DNA integrity and stability. Dysregulation of CKI gamma2 has been linked to various diseases, including cancer and neurodegenerative disorders, making it a valuable biomarker for studying these conditions and developing novel treatment strategies.

With its user-friendly protocol and robust performance, the CKI Gamma2 Colorimetric Cell-Based ELISA Kit is the perfect tool for researchers looking to investigate the role of CKI gamma2 in cellular pathways and disease processes. Trust in this kit to deliver accurate and reliable data for your research needs.

Product Name:CKI-gamma2 Colorimetric Cell-Based ELISA
Product Code:CBCAB01050
ELISA Type:Cell-Based
Target:CKI-gamma2
Reactivity:Human, Mouse, Rat
Dynamic Range:> 5000 Cells
Detection Method:Colorimetric 450 nmStorage/Stability:4°C/6 Months
Format:96-Well Microplate

The CKI-gamma2 Colorimetric Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can detect CKI-gamma2 protein expression profile in cells. The kit can be used for measuring the relative amounts of CKI-gamma2 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie siRNA or chemicals), or activators have on CKI-gamma2.

Qualitative determination of CKI-gamma2 concentration is achieved by an indirect ELISA format. In essence, CKI-gamma2 is captured by CKI-gamma2-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: 1455, UniProt ID: P78368, OMIM: 602214, Unigene: Hs.651905
Gene Symbol:CKI-gamma2
Sub Type:None
UniProt Protein Function:CK1G2: an ubiquitous protein kinase of the CK1 family. Involved in growth and morphogenesis. Two splice variant isoforms have been described.
UniProt Protein Details:

Protein type:EC 2.7.11.1; Kinase, protein; Protein kinase, Ser/Thr (non-receptor); Protein kinase, CK1; CK1 group; CK1 family

Chromosomal Location of Human Ortholog: 19p13.3

Cellular Component: membrane; cytosol; nucleus

Molecular Function:protein serine/threonine kinase activity; protein binding; magnesium ion binding; phosphoprotein binding; glycoprotein binding; peptide binding; ATP binding

Biological Process: regulation of cell shape; peptidyl-serine phosphorylation; sphingolipid metabolic process; Wnt receptor signaling pathway; sphingolipid biosynthetic process; protein amino acid autophosphorylation; endocytosis; signal transduction; protein amino acid phosphorylation

UniProt Code:P78368
NCBI GenInfo Identifier:3024060
NCBI Gene ID:1455
NCBI Accession:P78368.1
UniProt Secondary Accession:P78368,O00704, Q8WUB1, B5BU42,
UniProt Related Accession:P78368
Molecular Weight:415
NCBI Full Name:Casein kinase I isoform gamma-2
NCBI Synonym Full Names:casein kinase 1, gamma 2
NCBI Official Symbol:CSNK1G2  
NCBI Official Synonym Symbols:CK1g2  
NCBI Protein Information:casein kinase I isoform gamma-2; CKI-gamma 2; casein kinase 1 isoform gamma-2
UniProt Protein Name:Casein kinase I isoform gamma-2
Protein Family:Casein kinase
UniProt Gene Name:CSNK1G2  
UniProt Entry Name:KC1G2_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-CKI-gamma2 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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