Human GRIN2B (Glutamate Receptor, Ionotropic, N-Methyl-D-Aspartate 2B) ELISA Kit (HUES02633)

SKU:
HUES02633
€539

Description

ELISA Kit Technical ManualMSDS

Human GRIN2B (Glutamate Receptor, Ionotropic, N-Methyl-D-Aspartate 2B) 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 Human GRIN2B . 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 Human GRIN2B 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 Human GRIN2B, 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 Human GRIN2B. The concentration of Human GRIN2B in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityHuman
Detection MethodColormetric
Detection Range0.31-20 ng/mL
Sensitivity0.19 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human GRIN2B in samples. No significant cross-reactivity or interference between Human GRIN2B 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
202.363
2.367
2.3652.277
101.462
1.5
1.4811.393
50.946
0.906
0.9260.838
2.50.451
0.475
0.4630.375
1.250.262
0.244
0.2530.165
0.630.185
0.177
0.1810.093
0.310.132
0.14
0.1360.048
00.079
0.097
0.088--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human GRIN2B were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human GRIN2B were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)0.932.896.781.012.826.78
Standard deviation0.050.140.270.060.150.32
C V (%)5.384.843.985.945.324.72

Recovery

The recovery of Human GRIN2B 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)90-10196
EDTA plasma (n=5)87-10293
Cell culture media (n=5)91-10296

Linearity

Samples were spiked with high concentrations of Human GRIN2B 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 (%)91-10286-10185-96
Average (%)979290
1:4Range (%)89-10286-9889-102
Average (%)949294
1:8Range (%)93-10684-9582-93
Average (%)998988
1:16Range (%)87-10285-9988-104
Average (%)949095

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

Human GRIN2B (Glutamate Receptor, Ionotropic, N-Methyl-D-Aspartate 2B) ELISA Kit (HUES02633) 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.

Human GRIN2B (Glutamate Receptor, Ionotropic, N-Methyl-D-Aspartate 2B) ELISA Kit (HUES02633) Protein Information

UniProt Protein Function:NMDAR2B: an NMDA receptor subtype of glutamate-gated ion channels with high calcium permeability and voltage-dependent sensitivity to magnesium. Mediated by glycine. Plays a key role in synaptic plasticity, synaptogenesis, excitotoxicity, memory acquisition and learning. Mediates neuronal functions in glutamate neurotransmission. In concert with DAPK1 at extrasynaptic sites, acts as a central mediator for stroke damage. Its phosphorylation at Ser-1303 by DAPK1 enhances synaptic NMDA receptor channel activity inducing injurious Ca2+ influx through them, resulting in an irreversible neuronal death.
UniProt Protein Details:

Protein type:Channel, ligand-gated; Channel, calcium; Membrane protein, multi-pass; Membrane protein, integral

Chromosomal Location of Human Ortholog: 12p12

Cellular Component: postsynaptic membrane; synaptic vesicle; neuron projection; cell surface; integral to plasma membrane; dendrite; plasma membrane; cell junction; N-methyl-D-aspartate selective glutamate receptor complex

Molecular Function:protein binding; extracellular-glutamate-gated ion channel activity; zinc ion binding; glycine binding; N-methyl-D-aspartate selective glutamate receptor activity; calcium channel activity

Biological Process: synaptic transmission, glutamatergic; axon guidance; startle response; behavioral fear response; in utero embryonic development; glutamate signaling pathway; regulation of synaptic plasticity; learning; memory; synaptic transmission; detection of mechanical stimulus involved in sensory perception of pain; behavioral response to pain; sensory organ development; learning and/or memory; response to ethanol; suckling behavior; transport; ionotropic glutamate receptor signaling pathway; ephrin receptor signaling pathway; regulation of excitatory postsynaptic membrane potential

Disease: Mental Retardation, Autosomal Dominant 6; Epileptic Encephalopathy, Early Infantile, 27

NCBI Summary:N-methyl-D-aspartate (NMDA) receptors are a class of ionotropic glutamate receptors. NMDA receptor channel has been shown to be involved in long-term potentiation, an activity-dependent increase in the efficiency of synaptic transmission thought to underlie certain kinds of memory and learning. NMDA receptor channels are heteromers composed of three different subunits: NR1 (GRIN1), NR2 (GRIN2A, GRIN2B, GRIN2C, or GRIN2D) and NR3 (GRIN3A or GRIN3B). The NR2 subunit acts as the agonist binding site for glutamate. This receptor is the predominant excitatory neurotransmitter receptor in the mammalian brain. [provided by RefSeq, Jul 2008]
UniProt Code:Q13224
NCBI GenInfo Identifier:14548162
NCBI Gene ID:2904
NCBI Accession:Q13224.3
UniProt Secondary Accession:Q13224,Q12919, Q13220, Q13225, Q14CU4, Q9UM56,
UniProt Related Accession:Q13224
Molecular Weight:1484
NCBI Full Name:Glutamate receptor ionotropic, NMDA 2B
NCBI Synonym Full Names:glutamate receptor, ionotropic, N-methyl D-aspartate 2B
NCBI Official Symbol:GRIN2B
NCBI Official Synonym Symbols:MRD6; NR2B; hNR3; EIEE27; GluN2B; NMDAR2B
NCBI Protein Information:glutamate receptor ionotropic, NMDA 2B; NR3; glutamate receptor subunit epsilon-2; N-methyl-D-aspartate receptor subunit 3; N-methyl D-aspartate receptor subtype 2B; glutamate [NMDA] receptor subunit epsilon-2
UniProt Protein Name:Glutamate receptor ionotropic, NMDA 2B
UniProt Synonym Protein Names:Glutamate [NMDA] receptor subunit epsilon-2; N-methyl D-aspartate receptor subtype 2B; NMDAR2B; NR2B; N-methyl-D-aspartate receptor subunit 3; NR3; hNR3
Protein Family:Glutamate receptor ionotropic
UniProt Gene Name:GRIN2B
UniProt Entry Name:NMDE2_HUMAN
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Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
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