serum, plasma, tissue homogenates, lung lavage fluid and other biological fluids
Assay Length:
3.5h
Research Area:
Infection immunity;Pulmonology;
Uniprot ID:
P08427
Test principle:
The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Rat SPA. Standards or samples are added to the appropriate microtiter plate wells then with a biotin-conjugated antibody specific to Rat SPA. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Rat SPA, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of Rat SPA in the samples is then determined by comparing the OD of the samples to the standard curve.
標(biāo)準(zhǔn)曲線
Concentration (pg/mL)
OD
Corrected OD
1000.00
2.045
1.963
500.00
1.577
1.495
250.00
1.186
1.104
125.00
0.726
0.644
62.50
0.457
0.375
31.25
0.335
0.253
15.63
0.319
0.237
0.00
0.082
0.000
精密度
Intra-assay Precision (Precision within an assay):CV%<8%
Three samples of known concentration were tested twenty times on one plate to assess intra-assay precision.
Inter-assay Precision (Precision between assays):CV%<10%
Three samples of known concentration were tested in forty separate assays to assess inter-assay precision.
回收率
Matrices listed below were spiked with certain level of recombinant SPA and the recovery rates were calculated by comparing the measured value to the expected amount of SPA in samples.
Matrix
Recovery range
Average
serum(n=5)
80-96%
88%
EDTA plasma(n=5)
88-101%
94%
Heparin plasma(n=5)
78-91%
84%
線性
The linearity of the kit was assayed by testing samples spiked with appropriate concentration of SPA and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.