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Human E-Cad (E-Cadherin) ELISA Kit
Synonyms:CDH1; Arc-1; CD324; CDHE; LCAM; UVO; CAM 120/80; Epithelial Cadherin; Uvomorulin

Principle

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 E-Cad . 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 E-Cad and Avidin-Horseradish Peroxidase (HRP) conjugate is added to each micro plate well successively and then incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human E-Cad , biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color turns to 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 E-Cad . You can calculate the concentration of Human E-Cad in the samples by comparing the OD of the samples to the standard curve.

Product ID E-EL-H0014
Assay type Sandwich ELISA
Format 96T
Assay time 4.5h
Reactivity Human
Detection Method Colormetric
Detection Range 0.160—10.000ng/mL
Sensitivity 0.100ng/mL
Sample Volume Required Per Well 100μL
Sample Type serum, plasma and other biological fluids

Components

Item Specifications Storage
Micro ELISA Plate 8 wells ×12 strips 4℃/-20℃#
Reference Standard 2 vials 4℃/-20℃#
Reference Standard & Sample Diluent 1vial 20mL 4℃
Concentrated Biotinylated Detection Ab 1vial 120μL 4℃/-20℃#
Biotinylated Detection Ab Diluent 1vial 10mL 4℃
Concentrated HRP Conjugate 1vial 120μL 4℃(shading light)
HRP Conjugate Diluent 1vial 10mL 4℃
Concentrated Wash Buffer (25×) 1vial 30mL 4℃
Substrate Reagent 1vial 10mL 4℃(shading light)
 Stop Solution 1vial 10mL 4℃
Plate Sealer 5pieces
Manual 1 copy
Certificate of Analysis 1 copy
#: keep the kit at 4℃ if it’s used within 30 days, keep at -20℃ for longer storage.

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Human E-Cad were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Human E-Cad 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)
0.500 1.100 4.900 0.500 1.000 4.900
Standard deviation 0.000 0.100 0.200 0.000 0.000 0.200
C V (%) 5.000 4.600 3.100 5.200 4.200 3.100

Recovery

The recovery of Human E-Cad spiked to three different levels in samples throughout the range of the assay in various matrices was evaluated.

Sample Type Range (%) Average Recovery (%)
Serum (n=5) 88-105 96.000
EDTA plasma (n=5) 94-109 101.000
Cell culture media (n=5) 89-100 95.000

Linearity

Samples were spiked with high concentrations of Human E-Cad 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)
1:2 Range (%) 92-103 92-105 85-99
Average (%) 98.000 97.000 91.000
1:4 Range (%) 101-116 84-97 96-112
Average (%) 107.000 90.000 102.000
1:8 Range (%) 101-112 84-99 91-107
Average (%) 106.000 90.000 98.000
1:16 Range (%) 95-110 85-100 98-112
Average (%) 101.000 91.000 105.000

Specificity

This kit recognizes recombinant and natural Human E-Cad . No significant cross-reactivity or interference was observed.

Procedure

  1. Add 100μL standard or sample to each well. Incubate 90 mintues at 37℃.
  2. Add 100μL Biotinylated Detection Ab. Incubate 1 hour at 37℃.
  3. Aspirate and wash 3 times.
  4. Add 100μL HRP Conjugate. Incubate 30 minutes at 37℃.
  5. Aspirate and wash 5 times.
  6. Add 90μL Substrate Reagent. Incubate 15 minutes at 37℃.
  7. Add 50μL Stop Solution. Read it at 450nm immediately.
  8. Calculation of results.

Storage

All the reagents in the kit should be stored according to the labels on vials. Unused wells should be returned to the foil pouch with the desiccant pack and resealed along entire edge of zip-seal. Substrate Reagent shouldn't be kept at -20℃ (Check!). Exposure of reagents to strong light should be avoided in the process of incubation and storage. All the taps of reagents should be tightened to prevent evaporation and microbial contamination.

If you need the manual for more details, you can click here! 

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