Mouse CCND1 (Cyclin-D1) ELISA Kit (MOES00936)

SKU:
MOES00936
€549

Description

ELISA Kit Technical ManualMSDS

Mouse CCND1 (Cyclin-D1) ELISA Kit

This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Mouse CCND1 . Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse CCND1 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 CCND1, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Mouse CCND1. The concentration of Mouse CCND1 in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityMouse
Detection MethodColormetric
Detection Range0.16-10 ng/mL
Sensitivity0.10 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Mouse CCND1 in samples. No significant cross-reactivity or interference between Mouse CCND1 and analogues was observed.

Typical Data

As the OD 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
(ng/mL)
O.D Average Corrected
102.301
2.345
2.3232.27
51.617
1.643
1.631.577
2.50.863
0.863
0.8630.81
1.250.481
0.485
0.4830.43
0.630.226
0.224
0.2250.172
0.320.167
0.143
0.1550.102
0.160.097
0.113
0.1050.052
00.05
0.056
0.053--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse CCND1 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse CCND1 were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)0.521.594.410.481.474.26
Standard deviation0.040.100.230.030.080.15
C V (%)7.696.295.226.255.443.52

Recovery

The recovery of Mouse CCND1 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)94-10699
EDTA plasma (n=5)87-9992
Cell culture media (n=5)90-10596

Linearity

Samples were spiked with high concentrations of Mouse CCND1 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 (%)95-11197-11395-108
Average (%)103103101
1:4Range (%)92-10381-9383-95
Average (%)988689
1:8Range (%)91-10482-9284-98
Average (%)988790
1:16Range (%)86-9987-10088-99
Average (%)939293

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 ELISA 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 Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Mouse CCND1 (Cyclin-D1) ELISA Kit (MOES00936) 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 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
    11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
    12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.

Mouse CCND1 (Cyclin-D1) ELISA Kit (MOES00936) Protein Information

UniProt Protein Function:CCND1: a member of the highly conserved cyclin family, whose members are characterized by a dramatic periodicity in protein abundance through the cell cycle. Cyclins function as regulators of CDK kinases. Forms a complex with and functions as a regulatory subunit of CDK4 or CDK6, whose activity is required for cell cycle G1/S transition. This protein has been shown to interact with tumor suppressor protein Rb and the expression of this gene is regulated positively by Rb.
UniProt Protein Details:

Protein type:Nuclear receptor co-regulator; Activator; Cell cycle regulation; Oncoprotein

Cellular Component: nucleoplasm; tight junction; membrane; transcriptional repressor complex; cytoplasm; cyclin-dependent protein kinase holoenzyme complex; intracellular; nucleus

Molecular Function:protein binding; enzyme binding; histone deacetylase binding; protein complex binding; kinase activity; cyclin-dependent protein kinase regulator activity; transcription corepressor activity; transcription factor binding; protein kinase binding; protein kinase activity

Biological Process: lactation; fat cell differentiation; G1 DNA damage checkpoint; negative regulation of Wnt receptor signaling pathway; re-entry into mitotic cell cycle; positive regulation of cyclin-dependent protein kinase activity; regulation of cell cycle; positive regulation of mammary gland epithelial cell proliferation; negative regulation of epithelial cell differentiation; negative regulation of transcription from RNA polymerase II promoter; Wnt receptor signaling pathway through beta-catenin; protein amino acid phosphorylation; Leydig cell differentiation; regulation of transcription, DNA-dependent; positive regulation of cell proliferation; Wnt receptor signaling pathway; transcription, DNA-dependent; unfolded protein response; mammary gland epithelial cell proliferation; response to organic nitrogen; cell cycle; cell division; positive regulation of protein amino acid phosphorylation; regulation of cyclin-dependent protein kinase activity; regulation of protein kinase activity; response to DNA damage stimulus; G1/S transition of mitotic cell cycle

UniProt Code:P25322
NCBI GenInfo Identifier:116153
NCBI Gene ID:12443
NCBI Accession:P25322.1
UniProt Related Accession:P25322
Molecular Weight:36kDa
NCBI Full Name:G1/S-specific cyclin-D1
NCBI Synonym Full Names:cyclin D1
NCBI Official Symbol:Ccnd1  
NCBI Official Synonym Symbols:cD1; CycD1; Cyl-1; PRAD1; bcl-1; AI327039  
NCBI Protein Information:G1/S-specific cyclin-D1
UniProt Protein Name:G1/S-specific cyclin-D1
Protein Family:Chromosomal protein
UniProt Gene Name:Ccnd1  
UniProt Entry Name:CCND1_MOUSE
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
Sandwich
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