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ELK Biotechnology

ELK9777

Rat Phospho-Akt (S473) ELISA Kit

Rat Phospho-Akt (S473) ELISA Kit

Sensitivity

Rat Phospho-Akt (S473) ELISA Kit

Overview

Product name: Rat Phospho-Akt (S473) ELISA Kit
Reactivity: Rat
Alternative Names: p-AKT
Assay Type: Sandwich
Sensitivity: 6.7 pg/mL
Standard: 1000 pg/mL
Detection Range: 15.63-1000 pg/mL
Sample Type: tissue homogenates, cell lysates and other biological fluids
Assay Length: 3.5h
Research Area: Enzyme & Kinase;
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 Phospho-Akt. Standards or samples are added to the appropriate microtiter plate wells then with a biotin-conjugated antibody specific to Rat Phospho-Akt. 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 Phospho-Akt, 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 Phospho-Akt in the samples is then determined by comparing the OD of the samples to the standard curve.

Standard curve

Concentration (ng/mL) OD Corrected OD
1000.00 2.266 2.172
500.00 1.593 1.499
250.00 1.223 1.129
125.00 0.828 0.734
62.50 0.617 0.523
31.25 0.379 0.285
15.63 0.189 0.095
0.00 0.094 0.000

Precision

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.

Recovery

Matrices listed below were spiked with certain level of recombinant Phospho-Akt and the recovery rates were calculated by comparing the measured value to the expected amount of Phospho-Akt in samples.
Matrix Recovery range Average
serum(n=5) 96-107% 101%
EDTA plasma(n=5) 83-95% 89%
Heparin plasma(n=5) 86-99% 92%

Linearity

The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Phospho-Akt and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Matrix 1:2 1:4 1:8 1:16
serum(n=5) 91-99% 96-105% 95-103% 95-103%
EDTA plasma(n=5) 84-98% 93-102% 87-105% 83-92%
Heparin plasma(n=5) 82-96% 88-101% 90-99% 87-93%
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