Mouse S100A9 (S100 Calcium Binding Protein A9) CLIA Kit (MOES00609)

SKU:
MOES00609
€649

Description

ELISA Kit Technical ManualMSDS

Mouse S100A9 (S100 Calcium Binding Protein A9) CLIA Kit

This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Mouse S100A9 . Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Mouse S100A9 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 S100A9, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Mouse S100A9. The concentration of Mouse S100A9 in the samples can be calculated by comparing the RLU of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityMouse
Detection methodChemiluminescence
Detection range31.25-2000 pg/mL
Sensitivity18.75 pg/mL
Sample volume100uL
Sample typeSerum, plasma and other biological fluids
RepeatabilityCV < 15%

Specificity

This kit recognizes Mouse S100A9 in samples. No significant cross-reactivity or interference between Mouse S100A9 and analogues was observed.

Typical Data

As the RLU 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)
RLU Average Corrected
200054875
56567
5572155693
100022675
25273
2397423946
50011538
10526
1103211004
2504869
5719
52945266
1252662
2554
26082580
62.51416
1206
13111283
31.25666
682
674646
027
29
28--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Mouse S100A9 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Mouse S100A9 were tested on 3 different plates, 20 replicates in each plate.

  Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (pg/mL)97.12313.46998.0699.66287.411040.15
Standard deviation8.0525.4283.0411.7225.6166.99
C V (%)8.298.118.3211.768.916.44

Recovery

The recovery of Mouse S100A9 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)90-10696
EDTA plasma (n=5)83-9890
Cell culture media (n=5)93-10598

Linearity

Samples were spiked with high concentrations of Mouse S100A9 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 (%)86-9795-10992-106
Average (%)9210197
1:4Range (%)91-10584-9991-103
Average (%)999197
1:8Range (%)92-10489-10186-100
Average (%)979491
1:16Range (%)100-11890-10191-105
Average (%)1079697

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 CLIA 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 Reagent A1 vial, 5 mL4'C (shading light)
Substrate Reagent B1 vial, 5 mL4'C (shading light)
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Mouse S100A9 (S100 Calcium Binding Protein A9) CLIA Kit (MOES00609) 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 100 µL of standard solutions into the standard wells.
    3. Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100 µL 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 100 µL 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 350 µL 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 100 µL 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 100 µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37 °C. Protect the plate from light.
    11. Determine the RLU value of each well immediately.

Mouse S100A9 (S100 Calcium Binding Protein A9) CLIA Kit (MOES00609) Protein Information

UniProt Protein Function:S100A9: a calcium-binding regulatory protein of the S-100 family expressed by macrophages in acutely inflammated tissues and in chronic inflammations. May be an inhibitor of protein kinases. Also expressed in epithelial cells constitutively or induced during dermatoses. May interact with components of the intermediate filaments in monocytes and epithelial cells. Interacts with CEACAM3 in a calcium-dependent manner.
UniProt Protein Details:

Protein type:Calcium-binding

Cellular Component: cytoplasm; cytoskeleton; extracellular region; extracellular space; membrane; nucleus; plasma membrane

Molecular Function:antioxidant activity; arachidonic acid binding; calcium ion binding; metal ion binding; microtubule binding; RAGE receptor binding; zinc ion binding

Biological Process: actin cytoskeleton reorganization; apoptosis; astrocyte development; autophagy; caspase activation; chemotaxis; immune system process; inflammatory response; innate immune response; leukocyte chemotaxis; leukocyte migration during inflammatory response; neutrophil chemotaxis; peptidyl-cysteine S-nitrosylation; positive regulation of blood coagulation; positive regulation of inflammatory response; positive regulation of peptide secretion; regulation of inflammatory response; regulation of integrin biosynthetic process; regulation of translation

UniProt Code:P31725
NCBI GenInfo Identifier:399173
NCBI Gene ID:20202
NCBI Accession:P31725.3
UniProt Related Accession:P31725
Molecular Weight:13,049 Da
NCBI Full Name:Protein S100-A9
NCBI Synonym Full Names:S100 calcium binding protein A9 (calgranulin B)
NCBI Official Symbol:S100a9
NCBI Official Synonym Symbols:p14; Cagb; GAGB; L1Ag; BEE22; MRP14; 60B8Ag; AW546964
NCBI Protein Information:protein S100-A9
UniProt Protein Name:Protein S100-A9
UniProt Synonym Protein Names:Calgranulin-B; Leukocyte L1 complex heavy chain; Migration inhibitory factor-related protein 14; MRP-14; p14; S100 calcium-binding protein A9
Protein Family:Protein
UniProt Gene Name:S100a9
UniProt Entry Name:S10A9_MOUSE
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Additional Information

Product type:
ELISA
Reactivity:
Mouse
ELISA Type:
CLIA
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