Human VCAM-1/CD106 (Vascuolar Cell Adhesion Molecule 1) ELISA Kit (HUES03559)

SKU:
HUES03559
€299

Description

ELISA Kit Technical ManualMSDS

Human VCAM-1/CD106 (Vascuolar Cell Adhesion Molecule 1) 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 VCAM-1/CD106 . 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 VCAM-1/CD106 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 VCAM-1/CD106, 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 VCAM-1/CD106. The concentration of Human VCAM-1/CD106 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 Range1.56-100 ng/mL
Sensitivity0.94 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human VCAM-1/CD106 in samples. No significant cross-reactivity or interference between Human VCAM-1/CD106 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
(ng/mL)
O.D Average Corrected
1002.432
2.484
2.4582.403
501.63
1.642
1.6361.581
250.936
0.9
0.9180.863
12.50.421
0.433
0.4270.372
6.250.216
0.208
0.2120.157
3.130.162
0.144
0.1530.098
1.560.104
0.108
0.1060.051
00.051
0.059
0.055--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human VCAM-1/CD106 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human VCAM-1/CD106 were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)4.8211.4147.655.0811.9647.72
Standard deviation0.270.502.480.280.502.07
C V (%)5.604.385.205.514.184.34

Recovery

The recovery of Human VCAM-1/CD106 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)83-9890
EDTA plasma (n=5)94-105100
Cell culture media (n=5)94-110101

Linearity

Samples were spiked with high concentrations of Human VCAM-1/CD106 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 (%)94-10698-10990-107
Average (%)10010398
1:4Range (%)91-10285-9982-97
Average (%)979189
1:8Range (%)90-10582-9388-99
Average (%)968793
1:16Range (%)92-10582-9581-94
Average (%)998888

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 VCAM-1/CD106 (Vascuolar Cell Adhesion Molecule 1) ELISA Kit (HUES03559) 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 VCAM-1/CD106 (Vascuolar Cell Adhesion Molecule 1) ELISA Kit (HUES03559) Protein Information

UniProt Protein Function:VCAM1: Important in cell-cell recognition. Appears to function in leukocyte-endothelial cell adhesion. Interacts with the beta-1 integrin VLA4 on leukocytes, and mediates both adhesion and signal transduction. The VCAM1/VLA4 interaction may play a pathophysiologic role both in immune responses and in leukocyte emigration to sites of inflammation. 2 isoforms of the human protein are produced by alternative splicing.
UniProt Protein Details:

Protein type:Membrane protein, integral; Motility/polarity/chemotaxis; Cell adhesion

Chromosomal Location of Human Ortholog: 1p32-p31

Cellular Component: Golgi apparatus; extracellular space; cell surface; microvillus; endoplasmic reticulum; early endosome; integral to membrane; apical part of cell; plasma membrane; podosome; sarcolemma; filopodium; external side of plasma membrane

Molecular Function:amine oxidase activity; integrin binding; cell adhesion molecule binding

Biological Process: response to nicotine; regulation of immune response; chronic inflammatory response; extracellular matrix organization and biogenesis; viral reproduction; heart development; cytokine and chemokine mediated signaling pathway; cell-matrix adhesion; response to lipopolysaccharide; heterophilic cell adhesion; leukocyte adhesion; response to ethanol; response to zinc ion; B cell differentiation; membrane to membrane docking; response to hypoxia; positive regulation of T cell proliferation; amine metabolic process; response to ionizing radiation; cell adhesion; leukocyte tethering or rolling; acute inflammatory response; response to nutrient; aging

NCBI Summary:This gene is a member of the Ig superfamily and encodes a cell surface sialoglycoprotein expressed by cytokine-activated endothelium. This type I membrane protein mediates leukocyte-endothelial cell adhesion and signal transduction, and may play a role in the development of artherosclerosis and rheumatoid arthritis. Three alternatively spliced transcripts encoding different isoforms have been described for this gene. [provided by RefSeq, Dec 2010]
UniProt Code:P19320
NCBI GenInfo Identifier:137560
NCBI Gene ID:7412
NCBI Accession:P19320.1
UniProt Secondary Accession:P19320,Q6NUP8, A8K6R7, B4DKS4, E9PDD1,
UniProt Related Accession:P19320
Molecular Weight:74,346 Da
NCBI Full Name:Vascular cell adhesion protein 1
NCBI Synonym Full Names:vascular cell adhesion molecule 1
NCBI Official Symbol:VCAM1
NCBI Official Synonym Symbols:CD106; INCAM-100
NCBI Protein Information:vascular cell adhesion protein 1; CD106 antigen
UniProt Protein Name:Vascular cell adhesion protein 1
UniProt Synonym Protein Names:INCAM-100; CD_antigen: CD106
Protein Family:Vascular cell adhesion protein
UniProt Gene Name:VCAM1
UniProt Entry Name:VCAM1_HUMAN
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Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
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