ELISA Kit for Alpha-Fetoprotein (aFP)

A-FP; FETA; HPAFP; Alpha-Fetoglobulin; Alpha-1-fetoprotein


This assay has high sensitivity and excellent specificity for detection of Alpha-Fetoprotein (aFP).
No significant cross-reactivity or interference between Alpha-Fetoprotein (aFP) and analogues was observed.


Matrices listed below were spiked with certain level of recombinant Alpha-Fetoprotein (aFP) and the recovery rates were calculated by comparing the measured value to the expected amount of Alpha-Fetoprotein (aFP) in samples.

MatrixRecovery range (%)Average(%)
EDTA plasma(n=5)80-10181
heparin plasma(n=5)91-10498


Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level Alpha-Fetoprotein (aFP) were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level Alpha-Fetoprotein (aFP) were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV<10%
Inter-Assay: CV<12%


The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Alpha-Fetoprotein (aFP) and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.

EDTA plasma(n=5)81-99%82-99%96-104%81-90%
heparin plasma(n=5)88-105%80-90%95-102%98-105%


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

Pre-coated, ready to use 96-well strip plate1Plate sealer for 96 wells4
Standard2Standard Diluent1×20mL
Detection Reagent A1×120µLAssay Diluent A1×12mL
Detection Reagent B1×120µLAssay Diluent B1×12mL
TMB Substrate1×9mLStop Solution1×6mL
Wash Buffer (30 × concentrate)1×20mLInstruction manual1

Assay procedure summary

1. Prepare all reagents, samples and standards;
2. Add 100µL standard or sample to each well. Incubate 1 hours at 37°C;
3. Aspirate and add 100µL prepared Detection Reagent A. Incubate 1 hour at 37°C;
4. Aspirate and wash 3 times;
5. Add 100µL prepared Detection Reagent B. Incubate 30 minutes at 37°C;
6. Aspirate and wash 5 times;
7. Add 90µL Substrate Solution. Incubate 10-20 minutes at 37°C;
8. Add 50µL Stop Solution. Read at 450nm immediately.

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 Alpha-Fetoprotein (aFP). Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Alpha-Fetoprotein (aFP). 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 Alpha-Fetoprotein (aFP), 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 Alpha-Fetoprotein (aFP) in the samples is then determined by comparing the O.D. of the samples to the standard curve.



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