Rat NOS1/nNOS (Nitric Oxide Synthase 1, Neuronal) 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 Rat NOS1/nNOS . 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 Rat NOS1/nNOS 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 Rat NOS1/nNOS, 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 Rat NOS1/nNOS. The concentration of Rat NOS1/nNOS in samples can be calculated by comparing the OD of the samples to the standard curve.
|Detection Range||0.16-10 ng/mL|
|Sample Volume Required Per Well||100uL|
|Sample Type||Serum, plasma and other biological fluids|
This kit recognizes Rat NOS1/nNOS in samples. No significant cross-reactivity or interference between Rat NOS1/nNOS and analogues was observed.
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.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat NOS1/nNOS were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat NOS1/nNOS were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||6.00||4.38||4.22||6.00||5.71||3.19|
The recovery of Rat NOS1/nNOS spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||88-101||95|
|Cell culture media (n=5)||85-96||90|
Samples were spiked with high concentrations of Rat NOS1/nNOS 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)|
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.
|Micro ELISA Plate(Dismountable)||8 wells X12 strips||-20'C, 6 months|
|Reference Standard||2 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 Diluent||1 vial, 20 mL||4'C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25X)||1 vial, 30 mL|
|Substrate Reagent||1 vial, 10 mL||4'C(shading light)|
|Stop Solution||1 vial, 10 mL||4'C|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
Rat NOS1/nNOS (Nitric Oxide Synthase 1, Neuronal) ELISA Kit (RTES01008) 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.
Rat NOS1/nNOS (Nitric Oxide Synthase 1, Neuronal) ELISA Kit (RTES01008) Protein Information
|UniProt Protein Function:||nNOS: nitric oxide synthase 1, neuronal form. Produces nitric oxide which is a messenger molecule with diverse functions and displays many properties of a neurotransmitter in the brain and peripheral nervous system. May be an effector enzyme for the dystrophin complex. Four alternatively spliced isoforms have been described.|
|UniProt Protein Details:|
Protein type:EC 184.108.40.206; Amino Acid Metabolism - arginine and proline; Oxidoreductase
Cellular Component: azurophil granule; caveola; cytoplasm; cytoskeleton; cytosol; dendrite; dendritic spine; lipid raft; membrane; mitochondrial outer membrane; mitochondrion; nuclear membrane; nucleus; perinuclear region of cytoplasm; photoreceptor inner segment; plasma membrane; postsynaptic density; protein complex; sarcolemma; sarcoplasmic reticulum; sarcoplasmic reticulum membrane; synapse; T-tubule; vesicle membrane; Z disc
Molecular Function:amino acid binding; ATPase binding; cadmium ion binding; calmodulin binding; enzyme binding; FAD binding; FMN binding; heme binding; iron ion binding; NADP binding; nitric-oxide synthase activity; protein binding; protein homodimerization activity; sodium channel regulator activity
Biological Process: aging; arginine catabolic process; behavioral response to cocaine; exogenous drug catabolic process; female pregnancy; multicellular organismal response to stress; negative regulation of apoptosis; negative regulation of blood pressure; negative regulation of calcium ion transport; negative regulation of cell proliferation; negative regulation of heart contraction; negative regulation of hydrolase activity; negative regulation of insulin secretion; negative regulation of peptidyl-serine phosphorylation; negative regulation of potassium ion transport; negative regulation of serotonin uptake; negative regulation of vasoconstriction; nitric oxide biosynthetic process; nitric oxide mediated signal transduction; peptidyl-cysteine S-nitrosylation; positive regulation of guanylate cyclase activity; positive regulation of histone acetylation; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; positive regulation of vasodilation; reduction of cytosolic calcium ion concentration; regulation of heart contraction; regulation of neurogenesis; regulation of sensory perception of pain; regulation of sodium ion transport; response to activity; response to estrogen stimulus; response to ethanol; response to heat; response to hypoxia; response to lead ion; response to lipopolysaccharide; response to nicotine; response to nutrient levels; response to organic cyclic substance; response to organic nitrogen; response to peptide hormone stimulus; response to vitamin E; striated muscle contraction
|NCBI Summary:||enzyme that catalyzes the production of nitric oxide [RGD, Feb 2006]|
|NCBI GenInfo Identifier:||16258811|
|NCBI Gene ID:||24598|
|UniProt Secondary Accession:||P29476,P70594,|
|UniProt Related Accession:||P29476|
|Molecular Weight:||164,430 Da|
|NCBI Full Name:||nitric oxide synthase, brain|
|NCBI Synonym Full Names:||nitric oxide synthase 1|
|NCBI Official Symbol:||Nos1|
|NCBI Official Synonym Symbols:||bNOS|
|NCBI Protein Information:||nitric oxide synthase, brain|
|UniProt Protein Name:||Nitric oxide synthase, brain|
|UniProt Synonym Protein Names:||BNOS; Constitutive NOS; NC-NOS; NOS type I; Neuronal NOS; N-NOS; nNOS; Peptidyl-cysteine S-nitrosylase NOS1|
|Protein Family:||Nitric oxide synthase|
|UniProt Gene Name:||Nos1|
|UniProt Entry Name:||NOS1_RAT|