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Multiplex Assay Kit for Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,etc. by FLIA (Flow Luminescence Immunoassay)

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

Specificity

This assay has high sensitivity and excellent specificity for detection of Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,etc. by FLIA (Flow Luminescence Immunoassay).
No significant cross-reactivity or interference between Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,etc. by FLIA (Flow Luminescence Immunoassay) and analogues was observed.

Recovery

Matrices listed below were spiked with certain level of recombinant Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,etc. by FLIA (Flow Luminescence Immunoassay) and the recovery rates were calculated by comparing the measured value to the expected amount of Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,etc. by FLIA (Flow Luminescence Immunoassay) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 85-102 94
EDTA plasma(n=5) 86-94 91
heparin plasma(n=5) 83-102 96

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,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 Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,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 Mitochondrial Open Reading Frame Of The 12S rRNA-c (MOTS-c) ,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) 95-105% 96-104% 96-105% 78-101%
EDTA plasma(n=5) 92-99% 86-104% 82-92% 78-95%
heparin plasma(n=5) 88-95% 81-97% 94-101% 86-94%

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#:MOTS-c) 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 50μL standard or sample to each well,
    add 10μL magnetic beads,and 50μL Detection Reagent A,incubate 60min at 37°C on shaker;
3. Wash plate on magnetic frame for three times;
4. Add 100μL prepared Detection Reagent B, and incubate 30 min at 37°C on shaker;
5. Wash plate on magnetic frame for three times;
6. Add 100μL sheath solution, swirl for 2 minutes, read on the machine.

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Magazine Citations
Pediatric Diabetes  Circulating MOTS‐c levels are decreased in obese male children and adolescents and associated with insulin resistance Pubmed:29691953
Rapid Commun Mass Spectrom Development of a mass spectrometry‐based detection method for the mitochondrion‐derived peptide MOTS‐c in plasma samples for doping control purposes Pubmed: 30394592
DIABETOLOGIA Adiponectin treatment improves insulin resistance in mice by regulating the expression of the mitochondrial-derived peptide MOTS-c and its response to exercise via … Pubmed: 32880686
Induction of hyperandrogenism and insulin resistance differentially modulates ferroptosis in uterine and placental tissues of pregnant rats
FASEB JOURNAL Neural cell©\derived plasma exosome protein abnormalities implicate mitochondrial impairment in first episodes of psychosis 33454965
Biochim Biophys Acta Mol Basis Dis MOTS-c interacts synergistically with exercise intervention to regulate PGC-1¦Á expression, attenuate insulin resistance and enhance glucose metabolism in mice via?¡­ 33722744
researchsquare Evolution of Mitochondrial-Derived Peptides, Humanin and MOTSc and Changes in Insulin Sensitivity During Early Gestation
Mol Psychiatry Abnormal levels of mitochondrial proteins in plasma neuronal extracellular vesicles in major depressive disorder 34471251
Pharmacol Res The Mitochondrial-Derived Peptide MOTS-c Relieves Hyperglycemia and Insulin Resistance in Gestational Diabetes Mellitus 34798268
Ann Neurol SARS‐CoV‐2 and mitochondrial proteins in neural‐derived exosomes of COVID‐19 Pubmed:35285072
J Clin Med Evolution of Mitochondrially Derived Peptides Humanin and MOTSc, and Changes in Insulin Sensitivity during Early Gestation in Women with and without Gestational … Pubmed:35683389
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