Multiplex Assay Kit for S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay)

MRP8; MIF; 60B8AG; CAGA; CFAG; CGLA; CP-10; L1Ag; NIF; P8; Calgranulin A; Cystic fibrosis antigen; Leukocyte L1 complex light chain; Migration inhibitory factor-related protein 8

(Note: Up to 8-plex in one testing reaction)

Specificity

This assay has high sensitivity and excellent specificity for detection of S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay).
No significant cross-reactivity or interference between S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) and analogues was observed.

Recovery

Matrices listed below were spiked with certain level of recombinant S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) and the recovery rates were calculated by comparing the measured value to the expected amount of S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 94-102 99
EDTA plasma(n=5) 88-97 92
heparin plasma(n=5) 89-97 92
sodium citrate plasma(n=5) 95-105 99

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%

Linearity

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of S100 Calcium Binding Protein A8 (S100A8) ,etc. by FLIA (Flow Luminescence Immunoassay) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

Sample 1:2 1:4 1:8 1:16
serum(n=5) 98-105% 87-95% 88-95% 95-104%
EDTA plasma(n=5) 81-96% 92-101% 83-97% 80-101%
heparin plasma(n=5) 81-93% 84-102% 87-103% 92-105%
sodium citrate plasma(n=5) 95-102% 82-104% 78-96% 84-101%

Stability

The stability of kit is determined by the loss rate of activity. The loss rate of this kit is less than 5% within the expiration date under appropriate storage condition.
To minimize extra influence on the performance, operation procedures and lab conditions, especially room temperature, air humidity, incubator temperature should be strictly controlled. It is also strongly suggested that the whole assay is performed by the same operator from the beginning to the end.

Reagents and materials provided

Reagents Quantity Reagents Quantity
96-well plate 1 Plate sealer for 96 wells 4
Pre-Mixed Standard 2 Standard Diluent 1×20mL
Pre-Mixed Magnetic beads (22#:S100A8) 1 Analysis buffer 1×20mL
Pre-Mixed Detection Reagent A 1×120μL Assay Diluent A 1×12mL
Detection Reagent B (PE-SA) 1×120μL Assay Diluent B 1×12mL
Sheath Fluid 1×10mL Wash Buffer (30 × concentrate) 1×20mL
Instruction manual 1

Assay procedure summary

1. Preparation of standards, reagents and samples before the experiment;
2. Add 100μL standard or sample to each well,
    add 10μL magnetic beads, and incubate 90min at 37°C on shaker;
3. Remove liquid on magnetic frame, add 100μL prepared Detection Reagent A. Incubate 60min at 37°C on shaker;
4. Wash plate on magnetic frame for three times;
5. Add 100μL prepared Detection Reagent B, and incubate 30 min at 37°C on shaker;
6. Wash plate on magnetic frame for three times;
7. Add 100μL sheath solution, swirl for 2 minutes, read on the machine.

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Magazine Citations
Biochemical and Biophysical Research Communications High circulating levels of S100A8/A9 complex (calprotectin) in male Japanese with abdominal adiposity and dysregulated expression of S100A8 and S100A9 in adipose tissues of obese mice ScienceDirect: S0006291X1200352X
American Journal of Reproductive Immunology Association of Increased S100A8 Serum Protein with Early Pregnancy Loss Pubmed:25252120
the faseb journal Soluble receptor for advanced glycation end products inhibits disease progression in autosomal dominant polycystic kidney disease by down-regulating cell proliferation PubMed: 25934702
J Immunol S100A8 Production in CXCR2-Expressing CD11b+ Gr-1high Cells Aggravates Hepatitis in Mice Fed a High-Fat and High-Cholesterol Diet PubMed: 26608915
PLoS One Analytical Performance of ELISA Assays in Urine: One More Bottleneck towards Biomarker Validation and Clinical Implementation Pubmed:26889680
Journal of Alzheimer's Disease Proteomics Analysis of Blood Serums from Alzheimer's Disease Patients Using iTRAQ LabelingTechnology. pubmed:27911324
 Journal of Functional Foods Standardized ethanolic extract of the rhizome of Curcuma xanthorrhiza prevents murine ulcerative colitis by regulation of inflammation S1756464617300208
International Journal of Molecular Medicine Proinflammatory effects of S100A8/A9 via TLR4 and RAGE signaling pathways in BV-2 microglial cells pubmed:28498464
International journal of molecular sciences Immune Modulating Topical S100A8/A9 Inhibits Growth of Pseudomonas aeruginosa and Mitigates Biofilm Infection in Chronic Wounds. pubmed:28672877
Gastroenterology Tu1938-Micronutrients Assessment in Patients with Cirrhosis 10.1016:S0016-5085(18)33544-3
Gastroenterology Tu1942-PI3K/AKT-Mediated Upregulation of C7 Inhibits Colorectal Cancer Proliferation and Metastasis 10.1016:S0016-5085(18)33544-5
Journal of Biological Chemistry Signaling between pancreatic β-cells and macrophages via S100 calcium-binding protein A8 exacerbates β-cell apoptosis and islet inflammation Pubmed:29496993
European review for medical and pharmacological sciences S100A8/A9 promotes MMP-9 expression in the fibroblasts from cardiac rupture after myocardial infarction by inducing macrophages secreting TNFα. Pubmed:29949169
Foodborne Pathogens and Disease Synergistic effect of immunomodulatory S100A8/A9 and ciprofloxacin against Pseudomonas aeruginosa biofilm in a murine chronic wound model Pubmed: 31116394
Kidney International Reports Systemic Inflammation Precedes Microalbuminuria in Diabetes
Cancer Management and Research iTRAQ-based quantitative protein expression profiling of biomarkers in childhood B-cell and T-cell acute lymphoblastic leukemia Pubmed: 31440093
scandinavian journal of clinical & laboratory investigation Serum S100A6, S100A8, S100A9 and S100A11 proteins in colorectal neoplasia: results of a single centre prospective study Pubmed: 31856598
ANNALS OF AGRICULTURAL AND ENVIRONMENTAL MEDICINE Analysis of novel serum markers of fibrosis and angiogenesis in patients with alcoholic liver cirrhosis
MOLECULAR VISION Quantification of a panel for dry-eye protein biomarkers in tears: A comparative pilot study using standard ELISA and customized microarrays 34012227
university of copenhagen Adjunctive S100A8/A9 Immunomodulation Hinders Cipro fl oxacin Resistance in
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