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Human NFKB-p65(Nuclear factor NF-kappa-B p65 subunit) ELISA Kit

Citations(2)Uniprot : Q04206

Cat.No.:E-EL-H1388
Reactivity: Human

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Product Details

Properties

Assay type	Sandwich-ELISA
Format	96T/48T
Assay time	3.5h
Detection range	0.16-10 ng/mL
Sensitivity	0.10 ng/mL
Sample type &Sample volume	serum, plasma and other biological fluids; 100μL
Specificity	This kit recognizes Human NFKB-p65 in samples. No significant cross-reactivity or interference between Human NFKB-p65 and analogues was observed.
Reproducibility	Both intra-CV and inter-CV are < 10%.
Application	This ELISA kit applies to the in vitro quantitative determination of Human NFKB-p65 concentrations in serum, plasma and other biological fluids.

Test Principle

This ELISA kit uses the Sandwich-ELISA principle. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human NFKB-p65. Standards or samples are added to the micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human NFKB-p65 and Avidin-Horseradish Peroxidase (HRP) conjugate are added successively to each micro plate well and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human NFKB-p65, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by the addition of 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 NFKB-p65. You can calculate the concentration of Human NFKB-p65 in the samples by comparing the OD of the samples to the standard curve.

Kit components & Storage

An unopened kit can be stored at 2-8℃ for 1 month. If the kit is not supposed to be used within 1 month, store the components separately according to the following conditions once the kit is received.

Item	Specifications	Storage
Micro ELISA Plate(Dismountable)	96T: 8 wells ×12 strips 48T: 8 wells ×6 strips	-20℃, 6 months
Reference Standard	96T: 2 vials 48T: 1 vial
Concentrated Biotinylated Detection Ab (100×)	96T: 1 vial, 120 μL 48T: 1 vial, 60 μL
Concentrated HRP Conjugate (100×)	96T: 1 vial, 120 μL 48T: 1 vial, 60 μL	-20℃(Protect from light), 6 months
Reference Standard & Sample Diluent	1 vial, 20 mL	2-8°C, 6 months
Biotinylated Detection Ab Diluent	1 vial, 14 mL
HRP Conjugate Diluent	1 vial, 14 mL
Concentrated Wash Buffer (25×)	1 vial, 30 mL
Substrate Reagent	1 vial, 10 mL	2-8℃(Protect from light)
Stop Solution	1 vial, 10 mL	2-8°C
Plate Sealer	5 pieces
Manual	1 copy
Certificate of Analysis	1 copy

Technical Data

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human NFKB-p65 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human NFKB-p65 were tested on 3 different plates, 20 replicates in each plate.

	Intra-assay Precision			Inter-assay Precision
Sample	1	2	3	1	2	3
n	20	20	20	20	20	20
Mean (ng/mL)	0.54	1.49	3.97	0.52	1.39	3.93
Standard deviation	0.03	0.06	0.21	0.03	0.07	0.13
CV (%)	5.56	4.03	5.29	5.77	5.04	3.31

Recovery

The recovery of Human NFKB-p65 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=8)	90-104	95
EDTA plasma (n=8)	92-105	100
Cell culture media (n=8)	86-97	92

Linearity

Samples were spiked with high concentrations of Human NFKB-p65 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

Target Information

Database Links	SwissProt: Q04206
Synonyms	NFKB3, p65, RelA
Research Area	Cancer, Cell Biology, Cardiovascular, Epigenetics and Nuclear Signaling, Metabolism, Microbiology, Neuroscience, Signal transduction

Assay Procedures

	1. Add 100μL standard or sample to the wells. Incubate for 90 min at 37°C
	2. Discard the liquid, immediately add 100μL Biotinylated Detection Ab working solution to each well. Incubate for 60 min at 37°C
	3. Aspirate and wash the plate for 3 times
	4. Add 100μL HRP conjugate working solution. Incubate for 30 min at 37°C. Aspirate and wash the plate for 5 times
	5. Add 90μL Substrate Reagent. Incubate for 15 min at 37°C
	6. Add 50μL Stop Solution
	7. Read the plate at 450nm immediately. Calculation of the results

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Citations

From now on, if you have published a paper by using any of our products since 1/1/2019, fill out the “Elabscience Publication Reward Application Form”carefully and send it to orders@elabscience.com, we will get back to you with the reward after we confirm it ASAP!

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Journal of Reproductive Immunology (2020) IF: 4.018

Inhibition of TLR2/TLR4 alleviates the Neisseria gonorrhoeae infection damage in human endometrial epithelial cells via Nrf2 and NF-Kβsignaling

DOI: 10.1016/j.jri.2020.103192
PMID: 32950783	Sample: Cell culture supernatant

Bioscience reports (2019) IF: 2.942

Up-regulated MCPIP1 in abdominal aortic aneurysm is associated with vascular smooth muscle cell apoptosis and MMPs production

DOI: 10.1042/BSR20191252
PMID: 31651935	Sample: Serum,Plasma,Cell sample

Reviews/Q&A

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Reviews
Q&A

K******************hSubmitted [ May 17 2022 ]

Asked: Dear/ Madam, we would like know about Human NFKB-p65(Nuclear factor NF-kappa-B p65 subunit) ELISA Kit.The antibody coated in Elisa plate are against which domain of p65? This kit will give total nf-kb content in the cells or the activated nf-kb content?

adminSubmitted [ May 17 2022 ]

Answered: For this kit, the immunogen sequences for the capture and detection antibodies are recombinant human NFKB-p65 289-551(# Q04206), and the total NFKB-p65 content in the sample is detected.

T***********nSubmitted [ Mar 08 2021 ]

Asked: What are the epitopes of the two antibodies? I have a chimeric protein that has only the p65 activation domain that I am trying to assay. Are these antibodies polyclonal against the entire protein?

adminSubmitted [ Mar 08 2021 ]

Answered: The capture antibody is mouse monoclonal and the detection antibody is rabbit polyclonal, the immunogen is synthetic human NFKB-p65 aa 500 to the C-terminus for capture antibody and recombinant human NFKB-p65 aa 402-485 for detection antibody based on the sequence of # Q04206 in Uniprot.

D******************eSubmitted [ Jan 20 2019 ]

Asked: Dear colleagues, We would like to offer Human NF-κB p65(Nuclear Factor Kappa B p65) ELISA Kit. How can we do it?

adminSubmitted [ Jan 20 2019 ]

Answered: Dear Dr Natalia Chebotare, Thanks for your interest in our product? This is Cathy Mao for your service. Can you advise where are you from? The catalogue number for the kit inquired is E-EL-H1388, retail price is $580, usually we will charge $50 per order as freight cost which will be under our DHL or FedEx account, if you have your own express, this cost will be free of charge for you. Lead time is 3 working days and delivery will be made upon receipt of payment or bank slip, shelf life of our kit is 6 months. If you have any other question, please feel free to ask me. Thanks and have a nice day~ Regards, Cathy

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