Multiplex Assay Kit for Angiotensin I Converting Enzyme 2 (ACE2) ,etc. by FLIA (Flow Luminescence Immunoassay)

ACEH; ACEII; ACE-II; Peptidyl-Dipeptidase A; ACE-related carboxypeptidase; Angiotensin-converting enzyme homolog; Metalloprotease MPROT15

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

Specificity

This assay has high sensitivity and excellent specificity for detection of Angiotensin I Converting Enzyme 2 (ACE2) ,etc. by FLIA (Flow Luminescence Immunoassay).
No significant cross-reactivity or interference between Angiotensin I Converting Enzyme 2 (ACE2) ,etc. by FLIA (Flow Luminescence Immunoassay) and analogues was observed.

Recovery

Matrices listed below were spiked with certain level of recombinant Angiotensin I Converting Enzyme 2 (ACE2) ,etc. by FLIA (Flow Luminescence Immunoassay) and the recovery rates were calculated by comparing the measured value to the expected amount of Angiotensin I Converting Enzyme 2 (ACE2) ,etc. by FLIA (Flow Luminescence Immunoassay) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 78-96 92
EDTA plasma(n=5) 82-94 88
heparin plasma(n=5) 98-105 102

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Angiotensin I Converting Enzyme 2 (ACE2) ,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 Angiotensin I Converting Enzyme 2 (ACE2) ,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 Angiotensin I Converting Enzyme 2 (ACE2) ,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% 87-103% 86-95% 85-95%
EDTA plasma(n=5) 94-103% 81-101% 86-102% 93-101%
heparin plasma(n=5) 83-98% 89-96% 79-91% 95-104%

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#:ACE2) 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
Nature Medicine Angiotensin-Converting Enzyme 2 Overexpression Remarkably Ameliorated Glomerular Injury in a Rat Model of Diabetic Nephropathy: A Comparison with ACE Inhibition MolMed: 10_11_liu
Journal of Cancer Therapy Plasma Levels of Angiotensin-Converting Enzymes 1 and 2 and AGTR2 (T1247G and A5235G) Gene Polymorphisms Are Associated to Breast Cancer Progression. Scirp: Source
Reprod Toxicol.? ACE2 activation by xanthenone prevents leptin-induced increases in blood pressure and proteinuria during pregnancy in Sprague-Dawley rats Pubmed:25205467
Intensive Care Medicine Experimental Angiotensin converting enzymes in patients with acute respiratory distress syndrome Content: 3
Journal of Chromatography A Application of a new procedure for liquid chromatography/mass spectrometry profiling of plasma amino acid-related metabolites and untargeted shotgun proteomics to identify mechanisms and biomarkers of calcific aortic stenosis S0021967317311858
Journal of Clinical Pharmacy and Therapeutics Relationship between genetic variants of ACE2 gene and circulating levels of ACE2 and its metabolites. pubmed:28895159
Innate Immunity  Lipoxin A attenuates LPS-induced acute lung injury via activation of the ACE2-Ang-(1-7)-Mas axis Pubmed:29969931
Journal of Surgical Research Impact of angiotensin-converting enzyme 2 levels on postoperative pneumonia after esophagectomy Pubmed:29506841
canadian journal of physiology and pharmacology Could cardioprotective effect of ACE2 activator “diminazene aceturate” is more potent than ACE inhibitor “Enalapril” on acute myocardial infarction in rats? Pubmed: 30840489
Intensive Care Medicine Angiotensin-converting enzymes in acute respiratory distress syndrome
BioMed Research International AVE 0991 Attenuates Pyroptosis and Liver Damage after Heatstroke by Inhibiting the ROS-NLRP3 Inflammatory Signalling Pathway
Heart and vessels Relationship between circulating levels of angiotensin-converting enzyme 2-angiotensin-(1–7)-MAS axis and coronary heart disease Pubmed: 31359146
Medicine Alteration and association between serum ACE2/angiotensin (1-7)/Mas axis and oxidative stress in chronic kidney disease: A pilot study Pubmed: 32756181
American Journal of Respiratory and Critical Care Medicine Equilibrium Angiotensin Metabolite Profiling in Patients with Acute Respiratory Distress Syndrome Indicates Angiotensin Converting Enzyme Inhibition Pubmed: 32628511
Clinical Research in Cardiology Determinants of soluble angiotensin-converting enzyme 2 concentrations in adult patients with complex congenital heart disease 33280062
Clinical & Translational Immunology Pattern of circulating SARS©\CoV©\2©\specific antibody©\secreting and memory B©\cell generation in patients with acute COVID©\19 33552508
researchsquare Novel transgenic mice with Cre-dependent co-expression of GFP and human ACE2: a safe tool for study of COVID-19 pathogenesis 33855640
Viruses Different Neutralization Sensitivity of SARS-CoV-2 Cell-to-Cell and Cell-Free Modes of Infection to Convalescent Sera
Sci Rep Immunoreactivity of the SARS-CoV-2 entry proteins ACE-2 and TMPRSS-2 in murine models of hormonal manipulation, ageing, and cardiac injury 34907257
European Journal of Clinical Investigation Angiotensin‐converting enzyme 2 and Transmembrane protease serine 2 in female and male patients with end‐stage kidney disease Pubmed:35366343
Metabolites Ramipril Reduces Acylcarnitines and Distinctly Increases Angiotensin-Converting Enzyme 2 Expression in Lungs of Rats Pubmed:35448480
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