Mouse ATM (Ataxia Telangiectasia Mutated) ELISA Kit (MOES00747)

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MOES00747
€549

Description

ELISA Kit Technical ManualMSDS

Mouse ATM (Ataxia Telangiectasia Mutated) 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 Mouse ATM . 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 Mouse ATM 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 Mouse ATM, 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 Mouse ATM. The concentration of Mouse ATM in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityMouse
Detection MethodColormetric
Detection Range7.81-500 pg/mL
Sensitivity4.69 pg/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Mouse ATM in samples. No significant cross-reactivity or interference between Mouse ATM 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
5002.403
2.439
2.4212.352
2501.595
1.631
1.6131.544
1251.001
0.973
0.9870.918
62.50.487
0.489
0.4880.419
31.250.262
0.24
0.2510.182
15.630.177
0.167
0.1720.103
7.810.115
0.129
0.1220.053
00.065
0.073
0.069--

Precision

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

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

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)23.2465.14166.6821.0061.79169.25
Standard deviation1.583.207.881.333.195.13
C V (%)6.804.914.736.335.163.03

Recovery

The recovery of Mouse ATM 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)83-9690
EDTA plasma (n=5)93-110101
Cell culture media (n=5)86-10092

Linearity

Samples were spiked with high concentrations of Mouse ATM 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 (%)91-10388-10086-97
Average (%)979592
1:4Range (%)91-10483-9481-96
Average (%)968888
1:8Range (%)90-10383-9582-96
Average (%)958987
1:16Range (%)90-10286-10085-95
Average (%)959389

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

Mouse ATM (Ataxia Telangiectasia Mutated) ELISA Kit (MOES00747) 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.

Mouse ATM (Ataxia Telangiectasia Mutated) ELISA Kit (MOES00747) Protein Information

UniProt Protein Function:Serine/threonine protein kinase which activates checkpoint signaling upon double strand breaks (DSBs), apoptosis and genotoxic stresses such as ionizing ultraviolet A light (UVA), thereby acting as a DNA damage sensor. Recognizes the substrate consensus sequence [ST]-Q. Phosphorylates 'Ser-139' of histone variant H2AX/H2AFX at double strand breaks (DSBs), thereby regulating DNA damage response mechanism. Also plays a role in pre-B cell allelic exclusion, a process leading to expression of a single immunoglobulin heavy chain allele to enforce clonality and monospecific recognition by the B-cell antigen receptor (BCR) expressed on individual B-lymphocytes. After the introduction of DNA breaks by the RAG complex on one immunoglobulin allele, acts by mediating a repositioning of the second allele to pericentromeric heterochromatin, preventing accessibility to the RAG complex and recombination of the second allele. Also involved in signal transduction and cell cycle control. May function as a tumor suppressor. Necessary for activation of ABL1 and SAPK. Phosphorylates DYRK2, CHEK2, p53/TP53, FANCD2, NFKBIA, BRCA1, CTIP, nibrin (NBN), TERF1, RAD9 and DCLRE1C. May play a role in vesicle and/or protein transport. Could play a role in T-cell development, gonad and neurological function. Binds DNA ends. Plays a role in replication-dependent histone mRNA degradation. Phosphorylation of DYRK2 in nucleus in response to genotoxic stress prevents its MDM2-mediated ubiquitination and subsequent proteasome degradation. Phosphorylates ATF2 which stimulates its function in DNA damage response.
UniProt Code:Q62388
NCBI GenInfo Identifier:163838660
NCBI Gene ID:11920
NCBI Accession:NP_031525.2
UniProt Secondary Accession:Q62388,E9QNY7,
UniProt Related Accession:Q62388
Molecular Weight:349,418 Da
NCBI Full Name:serine-protein kinase ATM
NCBI Synonym Full Names:ataxia telangiectasia mutated
NCBI Official Symbol:Atm  
NCBI Official Synonym Symbols:AI256621; C030026E19Rik  
NCBI Protein Information:serine-protein kinase ATM
UniProt Protein Name:Serine-protein kinase ATM
UniProt Synonym Protein Names:Ataxia telangiectasia mutated homolog; A-T mutated homolog
Protein Family:Serine-protein kinase
UniProt Gene Name:Atm  
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Additional Information

Reactivity:
Mouse
ELISA Type:
Sandwich
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