Mouse GDN(Glia Derived Nexin)ELISA Kit  (MOES01783)

SKU:
MOES01783
€549

Description

ELISA Kit Technical ManualMSDS

Mouse GDN(Glia Derived Nexin)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 GDN . 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 GDN 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 GDN, 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 GDN. The concentration of Mouse GDN 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 Range6.25-400 pg/mL
Sensitivity3.75 pg/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Mouse GDN in samples. No significant cross-reactivity or interference between Mouse GDN 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
4002.526
2.558
2.5422.461
2001.713
1.729
1.7211.64
1001.013
1.009
1.0110.93
500.483
0.515
0.4990.418
250.273
0.267
0.270.189
12.50.188
0.182
0.1850.104
6.250.128
0.14
0.1340.053
00.072
0.09
0.081--

Precision

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

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

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)18.4451.91198.9017.0951.98204.75
Standard deviation0.952.2110.501.022.916.61
C V (%)5.154.265.285.975.603.23

Recovery

The recovery of Mouse GDN 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)88-10093
EDTA plasma (n=5)89-10296
Cell culture media (n=5)87-9994

Linearity

Samples were spiked with high concentrations of Mouse GDN 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 (%)86-9697-11085-100
Average (%)9110391
1:4Range (%)92-10685-9785-96
Average (%)999091
1:8Range (%)92-10785-9984-98
Average (%)999190
1:16Range (%)92-10878-9286-98
Average (%)998593

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 GDN(Glia Derived Nexin)ELISA Kit  (MOES01783) 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 GDN(Glia Derived Nexin)ELISA Kit  (MOES01783) Protein Information

UniProt Protein Function:SERPINE2: Serine protease inhibitor with activity toward thrombin, trypsin, and urokinase. Promotes neurite extension by inhibiting thrombin. Binds heparin. Belongs to the serpin family. 3 isoforms of the human protein are produced by alternative splicing.
UniProt Protein Details:

Protein type:Secreted; Inhibitor; Secreted, signal peptide

Cellular Component: extracellular matrix; extracellular space; extrinsic to external side of plasma membrane; membrane; cell soma; cell; extracellular region; cytosol; neuromuscular junction; vesicle

Molecular Function:serine-type endopeptidase inhibitor activity; heparin binding; glycosaminoglycan binding; protease inhibitor activity; receptor binding

Biological Process: negative regulation of proteolysis; nervous system development; regulation of synaptic transmission, glutamatergic; secretory granule organization and biogenesis; multicellular organismal development; cerebellar granular layer morphogenesis; negative regulation of peptidase activity; negative regulation of blood coagulation; mating plug formation; detection of mechanical stimulus involved in sensory perception; negative regulation of cell proliferation; negative regulation of smoothened signaling pathway; response to wounding; negative regulation of cell growth; blood coagulation; cell differentiation; negative regulation of protein catabolic process; regulation of timing of cell differentiation; negative regulation of phosphoinositide 3-kinase cascade; positive regulation of astrocyte differentiation; secretion by cell

UniProt Code:Q07235
NCBI GenInfo Identifier:6678099
NCBI Gene ID:20720
NCBI Accession:NP_033281.1
UniProt Secondary Accession:Q07235,Q921T7,
UniProt Related Accession:Q07235
Molecular Weight:44,207 Da
NCBI Full Name:glia-derived nexin
NCBI Synonym Full Names:serine (or cysteine) peptidase inhibitor, clade E, member 2
NCBI Official Symbol:Serpine2  
NCBI Official Synonym Symbols:PI7; PI-7; PN-1; Spi4; PAI-1; B230326M24Rik  
NCBI Protein Information:glia-derived nexin; GDN; serpin E2; protease nexin 1; protease nexin I; peptidase inhibitor 7; serine protease inhibitor 4; serine protease-inhibitor 4; serine (or cysteine) proteinase inhibitor, clade E, member 2; serine (or cysteine) proteinase inhibitor, clade E (nexin, plasminogen activator inhibitor type 1), member 2
UniProt Protein Name:Glia-derived nexin
UniProt Synonym Protein Names:Peptidase inhibitor 7; PI-7; Protease nexin 1; PN-1; Protease nexin I; Serine protease-inhibitor 4; Serpin E2
Protein Family:Glia-derived nexin
UniProt Gene Name:Serpine2  
UniProt Entry Name:GDN_MOUSE
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
Sandwich
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