Mouse CNTF (Ciliary Neurotrophic Factor) ELISA Kit (MOES00855)

SKU:
MOES00855
€549

Description

ELISA Kit Technical ManualMSDS

Mouse CNTF (Ciliary Neurotrophic Factor) 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 CNTF . 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 CNTF 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 CNTF, 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 CNTF. The concentration of Mouse CNTF 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 Range39.06-2500 pg/mL
Sensitivity23.44 pg/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Mouse CNTF in samples. No significant cross-reactivity or interference between Mouse CNTF 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
(pg/mL)
O.D Average Corrected
25002.323
2.343
2.3332.273
12501.407
1.453
1.431.37
6250.84
0.824
0.8320.772
312.50.35
0.382
0.3660.306
156.250.243
0.217
0.230.17
78.130.152
0.142
0.1470.087
39.060.101
0.111
0.1060.046
00.051
0.069
0.06--

Precision

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

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

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)115.09400.831183.56109.83373.091257.72
Standard deviation6.0121.9748.296.6022.0147.54
C V (%)5.225.484.086.015.903.78

Recovery

The recovery of Mouse CNTF 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)90-10698
EDTA plasma (n=5)93-10899
Cell culture media (n=5)87-9792

Linearity

Samples were spiked with high concentrations of Mouse CNTF 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 (%)97-11387-10285-97
Average (%)1039392
1:4Range (%)90-10478-9188-101
Average (%)968594
1:8Range (%)89-10283-9886-101
Average (%)958992
1:16Range (%)90-10582-9889-102
Average (%)978994

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 CNTF (Ciliary Neurotrophic Factor) ELISA Kit (MOES00855) 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 CNTF (Ciliary Neurotrophic Factor) ELISA Kit (MOES00855) Protein Information

UniProt Protein Function:CNTF: CNTF is a survival factor for various neuronal cell types. Seems to prevent the degeneration of motor axons after axotomy. Belongs to the CNTF family.
UniProt Protein Details:

Cellular Component: extracellular space; cell soma; axon; cytoplasm; nucleus; cytosol

Molecular Function:interleukin-6 receptor binding; growth factor activity; cytokine activity; ciliary neurotrophic factor receptor binding

Biological Process: negative regulation of photoreceptor cell differentiation; nervous system development; positive regulation of axon regeneration; protein heterooligomerization; multicellular organismal development; membrane hyperpolarization; positive regulation of hormone secretion; positive regulation of JAK-STAT cascade; JAK-STAT cascade; positive regulation of cell growth; positive regulation of tyrosine phosphorylation of Stat3 protein; positive regulation of phosphoinositide 3-kinase cascade; positive regulation of MAPKKK cascade; regulation of resting membrane potential; positive regulation of cell proliferation; neuron development; regulation of retinal cell programmed cell death; negative regulation of neuron apoptosis; cell differentiation; astrocyte activation; muscle morphogenesis; growth

NCBI Summary:The protein encoded by this gene is a polypeptide hormone whose actions appear to be restricted to the nervous system where it promotes neurotransmitter synthesis and neurite outgrowth in certain neuronal populations. The protein is a potent survival factor for neurons and oligodendrocytes, and it may be involved in reducing tissue destruction during inflammatory attacks. A read-through transcript variant composed of Zfp91 and Cntf sequences has been identified, but it is thought to be non-coding. Read-through transcription of Zfp91 and Cntf has been observed in both human and mouse. [provided by RefSeq, Aug 2008]
UniProt Code:P51642
NCBI GenInfo Identifier:1705963
NCBI Gene ID:12803
NCBI Accession:P51642.1
UniProt Related Accession:P51642
Molecular Weight:22,587 Da
NCBI Full Name:Ciliary neurotrophic factor
NCBI Synonym Full Names:ciliary neurotrophic factor
NCBI Official Symbol:Cntf  
NCBI Official Synonym Symbols:AI429687  
NCBI Protein Information:ciliary neurotrophic factor
UniProt Protein Name:Ciliary neurotrophic factor
Protein Family:Ciliary neurotrophic factor
UniProt Gene Name:Cntf  
UniProt Entry Name:CNTF_MOUSE
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
Sandwich
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