Human CVL/EZR (Cytovillin/Ezrin) ELISA Kit (HUES02990)

(No reviews yet) Write a Review
SKU:
HUES02990
€539

Description

ELISA Kit Technical ManualMSDS

Human CVL/EZR (Cytovillin/Ezrin) 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 Human CVL/EZR . 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 CVL/EZR 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 Human CVL/EZR, 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 Human CVL/EZR. The concentration of Human CVL/EZR in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityHuman
Detection MethodColormetric
Detection Range31.25-2000 pg/mL
Sensitivity18.75 pg/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human CVL/EZR in samples. No significant cross-reactivity or interference between Human CVL/EZR and analogues was observed.

Typical Data

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
(pg/mL)
O.D Average Corrected
20002.349
2.377
2.3632.297
10001.544
1.574
1.5591.493
5000.918
0.898
0.9080.842
2500.465
0.465
0.4650.399
1250.247
0.241
0.2440.178
62.50.177
0.155
0.1660.1
31.250.113
0.123
0.1180.052
00.058
0.074
0.066--

Precision

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

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human CVL/EZR were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)94.96291.52810.3786.93268.47805.77
Standard deviation6.5916.4426.744.8214.4436.42
C V (%)6.945.643.305.545.384.52

Recovery

The recovery of Human CVL/EZR 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=5)88-9993
EDTA plasma (n=5)88-10094
Cell culture media (n=5)88-10395

Linearity

Samples were spiked with high concentrations of Human CVL/EZR 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)
1:2Range (%)85-9894-11193-108
Average (%)92102101
1:4Range (%)87-10383-9588-98
Average (%)948893
1:8Range (%)92-10684-9685-96
Average (%)999091
1:16Range (%)92-10781-9385-99
Average (%)988792

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.

Item Specifications Storage
Micro ELISA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 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 Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Human CVL/EZR (Cytovillin/Ezrin) ELISA Kit (HUES02990) 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.

Human CVL/EZR (Cytovillin/Ezrin) ELISA Kit (HUES02990) Protein Information

UniProt Protein Function:Ezrin: a member of the ERM family which includes radixin and moesin. ERM proteins appear to function as cross-linkers between plasma membranes and actin-based cytoskeletons. Plays a key role in cell surface structure adhesion, migration, and organization.
UniProt Protein Details:

Protein type:Motility/polarity/chemotaxis; Cytoskeletal

Chromosomal Location of Human Ortholog: 6q25.3

Cellular Component: cortical cytoskeleton; extracellular space; microvillus; focal adhesion; basolateral plasma membrane; T-tubule; actin filament; cytosol; lipid raft; actin cytoskeleton; ruffle; extrinsic to membrane; membrane; apical part of cell; microvillus membrane; apical plasma membrane; nucleolus; plasma membrane; uropod; vesicle; filopodium

Molecular Function:actin filament binding; protein domain specific binding; protein binding; cell adhesion molecule binding

Biological Process: actin filament bundle formation; axon guidance; microvillus biogenesis; regulation of cell shape; leukocyte adhesion; filopodium formation; cytoskeletal anchoring; receptor internalization; membrane to membrane docking; establishment of epithelial cell polarity

NCBI Summary:The cytoplasmic peripheral membrane protein encoded by this gene functions as a protein-tyrosine kinase substrate in microvilli. As a member of the ERM protein family, this protein serves as an intermediate between the plasma membrane and the actin cytoskeleton. This protein plays a key role in cell surface structure adhesion, migration and organization, and it has been implicated in various human cancers. A pseudogene located on chromosome 3 has been identified for this gene. Alternatively spliced variants have also been described for this gene. [provided by RefSeq, Jul 2008]
UniProt Code:P15311
NCBI GenInfo Identifier:125987826
NCBI Gene ID:7430
NCBI Accession:P15311.4
UniProt Secondary Accession:P15311,P23714, Q4VX75, Q96CU8, Q9NSJ4, E1P5A8,
UniProt Related Accession:P15311
Molecular Weight:586
NCBI Full Name:Ezrin
NCBI Synonym Full Names:ezrin
NCBI Official Symbol:EZR  
NCBI Official Synonym Symbols:CVL; CVIL; VIL2; HEL-S-105  
NCBI Protein Information:ezrin; p81; villin-2; cytovillin 2; villin 2 (ezrin); epididymis secretory protein Li 105
UniProt Protein Name:Ezrin
UniProt Synonym Protein Names:Cytovillin; Villin-2; p81
Protein Family:Ezrin
UniProt Gene Name:EZR  
UniProt Entry Name:EZRI_HUMAN
View AllClose

Additional Information

Reactivity:
Human
ELISA Type:
Sandwich
View AllClose