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48 T
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Manual Download| Product Name | P5(Pregnenolone) ELISA Kit | Species | Universal |
| Uniprot ID | N/A | Alternative Names | Pregnenolone |
| Detection method | Competitive | Sensitivity | 0.47 ng/mL |
| Standard | 50 ng/mL | Detection Range | 0.78-50 ng/mL |
| Sample type | Serum, Plasma and Other biological fluids | ||
| Reaction time | 1.5H | Research Area | Reproductive science |
| Test principle | This ELISA kit uses the Competitive-ELISA principle. The micro ELISA plate provided in this kit has been pre-coated with P5. Standards or samples are added to the appropriate micro ELISA plate wells then with a biotin-conjugated antibody specific to P5. Next,Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added. 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 P5 in the samples is then determined by comparing the OD of the samples to the standard curve. | ||
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
| (ng/mL) | OD |
|---|---|
| 50.00 | 0.273 |
| 25.00 | 0.373 |
| 12.50 | 0.567 |
| 6.25 | 0.828 |
| 3.13 | 1.123 |
| 1.56 | 1.518 |
| 0.78 | 1.832 |
| 0.00 | 2.496 |
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level were tested on 3 different plates, 20 replicates in each plate, respectively.
| Intra-assay Precision | Inter-assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean(ng/mL) | 19.69 | 42.02 | 280.90 | 20.49 | 31.45 | 349.65 |
| Standard deviation | 2.10 | 4.64 | 14.51 | 2.61 | 5.77 | 18.06 |
| C V (%) | 5.12 | 7.11 | 4.59 | 3.20 | 6.73 | 4.22 |
The recovery of spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
| Sample Type | Range(%) | Average Recovery(%) |
|---|---|---|
| Serum (n=8) | 90-107 | 98 |
| EDTA plasma (n=8) | 87-99 | 93 |
| Cell culture media (n=8) | 82-98 | 90 |
Samples were spiked with high concentrations of target proteins and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
| Serum (n=5) | EDTA plasma (n=5) | Cell culture media (n=5) | ||
|---|---|---|---|---|
| 1:2 | Range (%) | 93-102 | 85-100 | 88-102 |
| Average (%) | 101 | 100 | 93 | |
| 1:4 | Range (%) | 90-99 | 85-100 | 86-93 |
| Average (%) | 92 | 93 | 88 | |
| 1:8 | Range (%) | 95-106 | 84-98 | 91-104 |
| Average (%) | 96 | 87 | 104 | |
| 1:16 | Range (%) | 87-101 | 95-101 | 87-98 |
| Average (%) | 94 | 99 | 89 |
