Human CD3g (Cluster of Differentiation 3g) ELISA Kit (HUES03615)

(No reviews yet) Write a Review
SKU:
HUES03615
€539

Description

ELISA Kit Technical ManualMSDS

Human CD3g (Cluster of Differentiation 3g) 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 CD3g . 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 CD3g 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 CD3g, 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 CD3g. The concentration of Human CD3g 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 Range0.16-10 ng/mL
Sensitivity0.10 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human CD3g in samples. No significant cross-reactivity or interference between Human CD3g 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
102.366
2.372
2.3692.278
51.556
1.602
1.5791.488
2.50.939
0.907
0.9230.832
1.250.485
0.523
0.5040.413
0.630.29
0.272
0.2810.19
0.320.204
0.176
0.190.099
0.160.138
0.146
0.1420.051
00.088
0.094
0.091--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human CD3g were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human CD3g were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)0.471.353.840.511.273.73
Standard deviation0.020.050.130.040.070.14
C V (%)4.263.703.397.845.513.75

Recovery

The recovery of Human CD3g 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)90-10597
EDTA plasma (n=5)94-109100
Cell culture media (n=5)88-9893

Linearity

Samples were spiked with high concentrations of Human CD3g 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 (%)95-10890-10186-102
Average (%)1029693
1:4Range (%)91-10486-10083-93
Average (%)989387
1:8Range (%)91-10482-9386-97
Average (%)968891
1:16Range (%)89-10582-9683-97
Average (%)978890

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 CD3g (Cluster of Differentiation 3g) ELISA Kit (HUES03615) 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 CD3g (Cluster of Differentiation 3g) ELISA Kit (HUES03615) Protein Information

UniProt Protein Function:Part of the TCR-CD3 complex present on T-lymphocyte cell surface that plays an essential role in adaptive immune response. When antigen presenting cells (APCs) activate T-cell receptor (TCR), TCR-mediated signals are transmitted across the cell membrane by the CD3 chains CD3D, CD3E, CD3G and CD3Z. All CD3 chains contain immunoreceptor tyrosine-based activation motifs (ITAMs) in their cytoplasmic domain. Upon TCR engagement, these motifs become phosphorylated by Src family protein tyrosine kinases LCK and FYN, resulting in the activation of downstream signaling pathways (PubMed:2470098). In addition to this role of signal transduction in T-cell activation, CD3G plays an essential role in the dynamic regulation of TCR expression at the cell surface (PubMed:8187769). Indeed, constitutive TCR cycling is dependent on the di-leucine-based (diL) receptor-sorting motif present in CD3G.
NCBI Summary:The protein encoded by this gene is the CD3-gamma polypeptide, which together with CD3-epsilon, -delta and -zeta, and the T-cell receptor alpha/beta and gamma/delta heterodimers, forms the T-cell receptor-CD3 complex. This complex plays an important role in coupling antigen recognition to several intracellular signal-transduction pathways. The genes encoding the epsilon, gamma and delta polypeptides are located in the same cluster on chromosome 11. Defects in this gene are associated with T cell immunodeficiency. [provided by RefSeq, Jul 2008]
UniProt Code:P09693
NCBI GenInfo Identifier:115993
NCBI Gene ID:917
NCBI Accession:P09693.1
UniProt Secondary Accession:P09693,Q2HIZ6,
UniProt Related Accession:P09693
Molecular Weight:20,469 Da
NCBI Full Name:T-cell surface glycoprotein CD3 gamma chain
NCBI Synonym Full Names:CD3g molecule
NCBI Official Symbol:CD3G  
NCBI Official Synonym Symbols:T3G; IMD17; CD3-GAMMA  
NCBI Protein Information:T-cell surface glycoprotein CD3 gamma chain
UniProt Protein Name:T-cell surface glycoprotein CD3 gamma chain
UniProt Synonym Protein Names:T-cell receptor T3 gamma chain; CD_antigen: CD3g
Protein Family:T-cell surface glycoprotein
UniProt Gene Name:CD3G  
View AllClose

Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
View AllClose

0 Reviews

View AllClose