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QuicKey Human NGAL(Neutrophil Gelatinase Associated Lipocalin) ELISA Kit

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Price:   $390
Cat Number:  E-TSEL-H0003
Lead Time:   7~10 days

Reactivity:  Human

Detection Range:  0.13~8 ng/mL

Sensitivity:  0.06 ng/mL

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Product Details

QuicKey Series

Get more sensitive and precise results with saving at least 1h comparing to traditional ELISA Kits. The new developed technology in house will help to accelerate your science research in a more efficient way.

Properties

Assay type Sandwich-ELISA
Format 96T/48T
Assay time 2.5 h
Reactivity Human
Detection method Colormetric
Colormetric 0.13-8 ng/mL
Sensitivity 0.06 ng/mL
Sample volume 50μL/well
Sample type serum, plasma, urine
Specificity This kit recognizes Human NGAL in samples. No significant cross-reactivity or interference between Human NGAL 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 NGAL concentrations in serum, plasma, urine. Please consult technical support for the applicability if other biological fluids need to be tested.

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 NGAL. Samples (or Standards) and biotinylated detection antibody specific for Human NGAL are added to the micro ELISA plate wells. Human NGAL would combine with the specific antibody. Then 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 NGAL, 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 ± 2 nm. The OD value is proportional to the concentration of Human NGAL. You can calculate the concentration of Human NGAL 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 six months. After test, the unused wells and reagents should be stored according to the table below. (The specific components might be slightly different in delivered kits, please refer to the manual for more information.)

Item Specifications Storage conditions after test
Micro ELISA Plate (Dismountable) 96T: 8 wells ×12 strips
48T: 8 wells ×6 strips
2-8℃, 1 month
Reference Standard 96T: 2 vials
48T: 1 vial
Discard unused reconstituted standard and dilutions
Reference Standard & Sample Diluent 1 vial, 20 mL 2-8℃
Biotinylated Detection Ab Working Solution 1 vial, 6 mL
HRP Conjugate Diluent 1 vial, 14 mL
Concentrated Wash Buffer (25×) 1 vial, 30 mL
Concentrated HRP Conjugate (100×) 96T: 1 vial, 120 μL
48T: 1 vial, 60 μL
2-8℃ (Protect from light)
Substrate Reagent 1 vial, 10 mL
Stop Solution 1 vial, 10 mL 2-8℃
Plate Sealer 5 pieces
Product Description 1 copy
Certificate of Analysis 1 copy

Note: All reagent bottle caps must be tightened to prevent evaporation and microbial pollution. The volume of reagents in partial shipments is a little more than the volume marked on the label, please use accurate measuring equipment instead of directly pouring into the vial(s).

Other supplies required

1. Microplate reader with 450nm wavelength filter

2. High-precision transfer pipette, EP tubes and disposable pipette tips

3. Incubator capable of maintaining 37℃

4. Deionized or distilled water

5. Absorbent paper

6. Loading slot for Wash Buffer

Technical Data

Sample values

sandwich-Ab-ELISA-Elabscience

Typical Standard Curve

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) 8 4 2 1 0.5 0.25 0.13 0
OD 2.569 2.13 1.533 1.088 0.656 0.437 0.225 0.059
Corrected OD 2.51 2.071 1.474 1.029 0.597 0.378 0.166 -
sandwich-Ab-ELISA-Elabscience

Precision

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

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human NGAL 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) 0.37 0.90 3.57 0.35 0.81 3.73
Standard deviation 0.02 0.04 0.16 0.02 0.04 0.17
CV (%) 5.41 4.44 4.48 5.71 4.94 4.56

Recovery

The recovery of Human NGAL 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) 86-100 94
EDTA plasma(n=8) 101-110 104
Urine(n=8) 87-98 90

Linearity

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

Serum(n=4) EDTA plasma (n=4) Urine(n=4)
1:2 Range (%) 88-99 90-101 101-110
Average (%) 91 96 104
1:4 Range (%) 95-105 92-102 96-109
Average (%) 99 97 102
1:8 Range (%) 87-96 86-100 98-111
Average (%) 92 93 105
1:16 Range (%) 101-111 90-98 100-110
Average (%) 105 95 104

Target information

Background

Neutrophil gelatinase associated lipocalin (NGAL) also known as Lipocalin 2 or Lcn2 is a 25 kDa, is a member of the lipocalin superfamily and a pleiotropic mediator of various inflammatory processes [1].NGAL, identified originally as a protein associated with matrix metalloproteinase 9 (MMP-9) of human neutrophils. Lipocalins are extracellular proteins which share a common tertiary structure that forms a barrel-like hydrophobic ligand binding site. When bound to MMP-9, NGAL protects it from proteolytic degradation sustaining the proteolytic activity of MMP-9. Neutrophil gelatinase-associated lipocalin (NGAL), is released by various cell types and is an attractive biomarker of inflammation, ischemia, infection, and kidney damage. Both intestinal and metabolic inflammation, as observed in obesity and related disorders, are associated with increased NGAL synthesis [2]. While NGAL in the intestinal tract regulates the composition of the gut microbiota and shows anti-inflammatory activities, it also exhibits proinflammatory activities in other experimental settings. Plasma NGAL is a novel marker of kidney function, which correlates to duration of end-stage renal failure (ESRD) and serum creatinine in uremic patients awaiting kidney transplantation. Plasma NGAL is associated with homocysteine in transplanted patients.
1. Kjeldsen L, Cowland J B, Borregaard N. Human neutrophil gelatinase-associated lipocalin and homologous proteins in rat and mouse.[J]. Biochim Biophys Acta, 2000, 1482(1):272-283.
2. Tilg, H., & Moschen, A. R. Adipocytokines: mediators linking adipose tissue, inflammation and immunity. Nature Reviews Immunology, 2006, 6(10), 772.

Database Links

SwissProt P80188 Human
Entrez Gene 3934 Human

Synonyms

LCN2, Lipocalin 2, Oncogene 24p3, MSFI

Documents

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Assay procedures

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