Multiplex Assay Kit for Protease Activated Receptor 2 (PAR2) ,etc. by FLIA (Flow Luminescence Immunoassay)

F2RL1; F2-RL1; GPR11; PAR2; Coagulation Factor II(thrombin)receptor-Like 1; G-protein coupled receptor 11; Thrombin receptor-like 1

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

Specificity

This assay has high sensitivity and excellent specificity for detection of Protease Activated Receptor 2 (PAR2) ,etc. by FLIA (Flow Luminescence Immunoassay).
No significant cross-reactivity or interference between Protease Activated Receptor 2 (PAR2) ,etc. by FLIA (Flow Luminescence Immunoassay) and analogues was observed.

Recovery

Matrices listed below were spiked with certain level of recombinant Protease Activated Receptor 2 (PAR2) ,etc. by FLIA (Flow Luminescence Immunoassay) and the recovery rates were calculated by comparing the measured value to the expected amount of Protease Activated Receptor 2 (PAR2) ,etc. by FLIA (Flow Luminescence Immunoassay) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 78-101 82
EDTA plasma(n=5) 98-105 102
heparin plasma(n=5) 93-101 98
sodium citrate plasma(n=5) 78-101 81

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Protease Activated Receptor 2 (PAR2) ,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 Protease Activated Receptor 2 (PAR2) ,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 Protease Activated Receptor 2 (PAR2) ,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) 86-104% 78-98% 83-93% 85-101%
EDTA plasma(n=5) 82-99% 86-95% 89-96% 88-95%
heparin plasma(n=5) 91-98% 78-95% 95-105% 90-104%
sodium citrate plasma(n=5) 97-104% 88-96% 78-95% 87-98%

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#:PAR2) 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
The Open Respiratory Medicine Journal Biomarkers of Fibroproliferative Healing in Fibrosing Idiopathic Interstitial Pneumonias PubMed: PMC3551240
Experimental Dermatology Cathepsin S, a new pruritus biomarker in clinical dandruff/seborrhoeic dermatitis evaluation Onlinelibrary: exd.12357
Exp Ther Med. PAR2, IL4R, TGFβ and TNFα in bronchoalveolar lavage distinguishes extrinsic allergic alveolitis from sarcoidosis Pubmed:Pmc4079423
BMC Neurol Proteinase-activated receptor 2 and disease biomarkers in cerebrospinal fluid in cases with autopsy-confirmed prion diseases and other neurodegenerative diseases PubMed: 25886404
Journal of Interferon & Cytokine Research Epithelial Cell-Derived Cytokines Contribute to the Pathophysiology of Eosinophilic Chronic Rhinosinusitis Pubmed:26540312
International journal of molecular sciences Cathepsin S Alters the Expression of Pro-Inflammatory Cytokines and MMP-9, Partially through Protease—Activated Receptor-2, in Human Corneal Epithelial Cells Pubmed: 30423938
Journal of Cosmetic Dermatology Whitening effects of cosmetic formulation in the vascular component of skin pigmentation Pubmed: 31074159
diagnostics Interstitial Score and Concentrations of IL-4R¦Á, PAR-2, and MMP-7 in Bronchoalveolar Lavage Fluid Could Be Useful Markers for Distinguishing Idiopathic Interstitial?¡­ 33924683
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