Multiplex Assay Kit for Calreticulin (CALR) ,etc. by FLIA (Flow Luminescence Immunoassay)

CRT; RO; SSA; CC1qR; ERp60; HACBP; grp60; CRTC; CRP55; Calregulin; Sicca Syndrome Antigen A; Autoantigen Ro; Endoplasmic reticulum resident protein 60

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

Specificity

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

Recovery

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

Matrix Recovery range (%) Average(%)
serum(n=5) 88-103 94
EDTA plasma(n=5) 86-94 90
heparin plasma(n=5) 90-97 94
sodium citrate plasma(n=5) 83-99 93

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Calreticulin (CALR) ,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 Calreticulin (CALR) ,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 Calreticulin (CALR) ,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) 91-98% 96-105% 86-101% 82-97%
EDTA plasma(n=5) 88-96% 96-104% 83-99% 78-99%
heparin plasma(n=5) 93-101% 78-91% 89-103% 92-105%
sodium citrate plasma(n=5) 86-101% 92-101% 82-99% 94-104%

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#:CALR) 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
Journal of Investigative Dermatology Oxidative Stress-Induced Calreticulin Expression and Translocation: New Insights into the Destruction of Melanocytes Pubmed: 23771121
International Journal of Molecular Sciences The Anticoagulant Effect of PGI2S and tPA in Transgenic Umbilical Vein Endothelial Cells Is Linked to Up-Regulation of PKA and PKC Mdpi: Source
The Journal of Maternal-Fetal & Neonatal Medicine Amniotic fluid calreticulin in pregnancies complicated by the preterm prelabor rupture of membranes Pubmed:26953684
journal of maternal-fetal & neonatal medicine Cervical fluid calreticulin and cathepsin-G in pregnancies complicated by preterm prelabor rupture of membranes. pubmed:28152632
Talanta Investigation of the interaction between calreticulin and immunoglobulin G by capillary electrophoresis and computer modeling Doi: 10.1016/j.talanta.2018.11.098
Sci Adv Thermostable ionizable lipid-like nanoparticle (iLAND) for RNAi treatment of hyperlipidemia Pubmed:35171673
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