Mouse CDKN1B (Cyclin Dependent Kinase Inhibitor 1B) CLIA Kit (MOES00211)

SKU:
MOES00211
€649

Description

ELISA Kit Technical ManualMSDS

Mouse CDKN1B (Cyclin Dependent Kinase Inhibitor 1B) CLIA Kit

This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Mouse CDKN1B . Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse CDKN1B 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 Mouse CDKN1B, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Mouse CDKN1B. The concentration of Mouse CDKN1B in the samples can be calculated by comparing the RLU of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityMouse
Detection methodChemiluminescence
Detection range3.13-200 pg/mL
Sensitivity1.88 pg/mL
Sample volume100uL
Sample typeSerum, plasma and other biological fluids
RepeatabilityCV < 15%

Specificity

This kit recognizes Mouse CDKN1B in samples. No significant cross-reactivity or interference between Mouse CDKN1B and analogues was observed.

Typical Data

As the RLU 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.

Concentration
(pg/mL)
RLU Average Corrected
20043867
44279
4407344035
10016783
19639
1821118173
508460
7952
82068168
253726
4144
39353897
12.51996
1968
19821944
6.251068
1036
10521014
3.13564
632
598560
037
39
38--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse CDKN1B were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse CDKN1B were tested on 3 different plates, 20 replicates in each plate.

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)9.1719.8494.208.4621.6788.66
Standard deviation0.891.4210.770.951.545.54
C V (%)9.717.1611.4311.237.116.25

Recovery

The recovery of Mouse CDKN1B spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)99-110105
EDTA plasma (n=5)93-10499
Cell culture media (n=5)86-9893

Linearity

Samples were spiked with high concentrations of Mouse CDKN1B 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)
1:2Range (%)102-11799-11288-102
Average (%)11010596
1:4Range (%)102-11787-10392-108
Average (%)1079499
1:8Range (%)100-11598-11585-101
Average (%)10710692
1:16Range (%)97-10993-10785-98
Average (%)10310091

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.

Item Specifications Storage
Micro CLIA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 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 Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent A1 vial, 5 mL4'C (shading light)
Substrate Reagent B1 vial, 5 mL4'C (shading light)
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Mouse CDKN1B (Cyclin Dependent Kinase Inhibitor 1B) CLIA Kit (MOES00211) 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 100µl of standard solutions into the standard wells.
    3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100µl 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 100µL 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 350µL 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 100µL 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 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
    11. Determine the RLU value of each well immediately.

Mouse CDKN1B (Cyclin Dependent Kinase Inhibitor 1B) CLIA Kit (MOES00211) Protein Information

UniProt Protein Function:p27Kip1: a cell-cycle regulatory protein that Interacts with cyclin-CDK2 and -CDK4, inhibiting cell cycle progression at G1. May mediate TGF beta-induced g1 arrest. Its degradation is triggered by its CDK dependent phosphorylation and subsequent ubiquitination by SCF complexes, is required for the cellular transition from quiescence to the proliferative state.
UniProt Protein Details:

Protein type:Cell cycle regulation; Oncoprotein; Protein kinase, regulatory subunit; Inhibitor

Cellular Component: cytoplasm; cytosol; nucleus; protein complex

Molecular Function:caspase activator activity; chaperone binding; cyclin-dependent protein kinase inhibitor activity; Hsp70 protein binding; protein binding; protein complex binding; protein phosphatase binding

Biological Process: autophagic cell death; caspase activation; cell cycle arrest; cell death; G1/S transition of mitotic cell cycle; inner ear development; negative regulation of apoptosis; negative regulation of cell growth; negative regulation of cell motility; negative regulation of cell proliferation; negative regulation of cyclin-dependent protein kinase activity; negative regulation of epithelial cell proliferation; negative regulation of kinase activity; negative regulation of mitotic cell cycle; negative regulation of phosphorylation; negative regulation of transcription, DNA-dependent; Notch signaling pathway; positive regulation of cell proliferation; positive regulation of cyclin-dependent protein kinase activity; positive regulation of microtubule polymerization; potassium ion transport; sensory perception of sound

UniProt Code:P46414
NCBI GenInfo Identifier:408360008
NCBI Gene ID:12576
NCBI Accession:P46414.2
UniProt Secondary Accession:P46414,Q8BG74,
UniProt Related Accession:P46414
Molecular Weight:22,193 Da
NCBI Full Name:Cyclin-dependent kinase inhibitor 1B
NCBI Synonym Full Names:cyclin-dependent kinase inhibitor 1B
NCBI Official Symbol:Cdkn1b  
NCBI Official Synonym Symbols:p27; Kip1; p27Kip1; AA408329; AI843786  
NCBI Protein Information:cyclin-dependent kinase inhibitor 1B
UniProt Protein Name:Cyclin-dependent kinase inhibitor 1B
UniProt Synonym Protein Names:Cyclin-dependent kinase inhibitor p27; p27Kip1
Protein Family:Cyclin-dependent kinase inhibitor
UniProt Gene Name:Cdkn1b  
UniProt Entry Name:CDN1B_MOUSE
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
CLIA
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