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Human HSP-70/HSPA9(Heat Shock 70 kDa Protein 9) ELISA Kit

Citations(5)Uniprot : P38646

Cat.No.:E-EL-H1863
Reactivity: Human

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Product Details

Properties

Assay type	Sandwich-ELISA
Format	96T/48T
Assay time	3.5h
Detection range	0.78-50 ng/mL
Sensitivity	0.47 ng/mL
Sample type &Sample volume	serum, plasma and other biological fluids; 100μL
Specificity	This kit recognizes Human HSP-70/HSPA9 in samples. No significant cross-reactivity or interference between Human HSP-70/HSPA9 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 HSP-70/HSPA9 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 HSP-70/HSPA9. 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 HSP-70/HSPA9 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 HSP-70/HSPA9, 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 HSP-70/HSPA9. You can calculate the concentration of Human HSP-70/HSPA9 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 HSP-70/HSPA9 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human HSP-70/HSPA9 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)	2.30	6.00	22.60	2.30	5.60	24.10
Standard deviation	0.10	0.30	1.10	0.10	0.30	0.90
CV (%)	4.35	5.00	4.87	4.35	5.36	3.73

Recovery

The recovery of Human HSP-70/HSPA9 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)	93-108	100
EDTA plasma (n=8)	91-106	97
Cell culture media (n=8)	91-103	98

Linearity

Samples were spiked with high concentrations of Human HSP-70/HSPA9 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

Target Information

Database Links	SwissProt: P38646
Synonyms	HSPA9, CSA, GRP-75, GRP75, HSPA9B, MOT, MOT2, MTHSP75, PBP74
Research Area	Cancer, Metabolism, 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

Citations

BIOSENSORS & BIOELECTRONICS (2018) IF: 8.173
A novel electrochemical immunosensor based on PG for early screening of depression markers-heat shock protein 70

DOI: 10.1016/j.bios.2018.03.049

Sample: Plasma
CANCER LETTERS (2020) IF: 7.36
Small extracellular vesicle-bound vascular endothelial growth factor secreted by carcinoma-associated fibroblasts promotes angiogenesis in a bevacizumab-resistant manner

DOI: 10.1016/j.canlet.2020.08.030

PMID: 32860852
Sample: Cell culture supernatant
Materials Science & Engineering C-Materials for Biological Applications (2020) IF: 5.88
Development of a sensitive electrochemical immunosensor using polyaniline functionalized graphene quantum dots for detecting a depression marker

DOI: 10.1016/j.msec.2020.110797

PMID: 32279828
Sample: Serum
ACS Omega (2022) IF: 3.512
Thymoquinone Induces Nrf2 Mediated Adaptive Homeostasis: Implication for Mercuric Chloride-Induced Nephrotoxicity

DOI: 10.1021/acsomega.2c00028

PMID: 35252727
Sample: liver
Annals of Translational Medicine (2022) IF: 3.932
Advanced platelet-rich fibrin promotes the paracrine function and proliferation of adipose-derived stem cells and contributes to micro-autologous fat transplantation by modulating HIF-1α and VEGF

DOI: 10.21037/atm-21-6812

Sample: adipose-derived stem cell (ASCs)

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Reviews/Q&A

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Reviews
Q&A

S**aSubmitted [ May 15 2019 ]

Asked: How can i dilute concentrated biotinylated detection Ab (100x) 14ml with biotinylated detection Ab diluent 120 microlitre?

adminSubmitted [ May 15 2019 ]

Answered: At first, the volume you mentioned is wrong for the two reagents, we provide 14mL biotinylated detection Ab diluent and 120 μL concentrated biotinylated detection Ab (100x) . The method to prepare biotinylated detection Ab working soltion： Calculate the required amount before the experiment (100 μL/well). In preparation, slightly more than calculated should be prepared. Centrifuge the stock tube before use, dilute the 100× Concentrated Biotinylated Detection Ab with Biotinylated Detection Ab Diluent at 1:99( eg. Mix 10μL concentrated biotinylated detection Ab(100x) and 990μL biotinylated detection Ab diluent to make 1mL biotinylated detection Ab working soltion).

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