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Multiplex Assay Kit for Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,etc. by FLIA (Flow Luminescence Immunoassay)

IHRP; PK120; H4P; ITIHL1; Plasma Kallikrein-Sensitive Glycoprotein; Plasma kallikrein sensitive glycoprotein 120; Inter-alpha-trypsin inhibitor family heavy chain-related protein

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

Specificity

This assay has high sensitivity and excellent specificity for detection of Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,etc. by FLIA (Flow Luminescence Immunoassay).
No significant cross-reactivity or interference between Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,etc. by FLIA (Flow Luminescence Immunoassay) and analogues was observed.

Recovery

Matrices listed below were spiked with certain level of recombinant Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,etc. by FLIA (Flow Luminescence Immunoassay) and the recovery rates were calculated by comparing the measured value to the expected amount of Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,etc. by FLIA (Flow Luminescence Immunoassay) in samples.

Matrix Recovery range (%) Average(%)
serum(n=5) 80-103 85
EDTA plasma(n=5) 85-97 91
heparin plasma(n=5) 98-105 102

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,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 Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,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 Inter Alpha-Globulin Inhibitor H4 (ITIH4) ,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% 91-102% 91-101% 89-96%
EDTA plasma(n=5) 92-99% 78-101% 81-98% 96-103%
heparin plasma(n=5) 92-99% 91-98% 79-101% 99-105%

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#:ITIH4) 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
Toxicology Letters Biomarkers for methotrexate-induced liver injury: urinary protein profiling of psoriasis patients. Pubmed: 23830989
International Journal of Medical Microbiology Proteomic demonstration of the recurrent presence of inter-alpha-inhibitor H4 heavy-chain during aspergillosis induced in an animal model Pubmed:24360996
Cancer Research and Treatment Inter-alpha Inhibitor H4 as a Potential Biomarker Predicting the Treatment Outcomes in Patients with Hepatocellular Carcinoma pubmed:28724284
Diagnostics Systemic Alterations of Immune Response-Related Proteins during Glaucoma Development in the Murine Model DBA/2J Pubmed: 32585848
Liver Cancer Biomarkers for locally advanced hepatocellular carcinoma patients treated with liver-directed combined radiotherapy
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