QuicKey Human TF(Transferrin) ELISA Kit
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| Price: | $390 |
Reactivity: Human
Detection Range: 0.14~100 ng/mL
Sensitivity: 0.13 ng/mL
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QuicKey Series
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Intended use
This ELISA kit applies to the in vitro quantitative determination of Human TF concentrations in serum, plasma, urine. Please consult technical support for the applicability if other biological fluids need to be tested.
Background
Transferrin is an iron binding protein found in blood plasma that is critical to cell health. Transferrin supplies iron to cells naturally in the body and as a supplement in cell culture media Transferrin (Tf) carries ferric iron from the intestine, reticuloendothelial system, and liver parenchymal cells to all proliferating cells in the body [1]. It shares homologous amino acid sequences with four other proteins: lactotransferrin, ovotransferrin, melanoma antigen p97, and HuBlym-1. Antigen p97 and the Tf receptor genes have been mapped on human chromosome 3. Transferrin has a molecular weight of around 80 kDa and contains two specific high-affinity Fe (III) binding sites. The affinity of transferrin for Fe (III) is extremely high (association constant is 1020M−1 at pH 7.4) but decreases progressively with decreasing pH below neutrality [2]. When not bound to iron, transferrin is known as Apo transferrin.
1. Sousa M D , Macedo M F . Transferrin and the Transferrin Receptor: Of Magic Bullets and Other Concerns [J]. Inflammation & Allergy Drug Targets, 2008, 7(1): 41-52.
2. Aisen P A , Leibman A , Zweier J L . Stoichiometric and site characteristics of the binding of iron to human transferrin [J]. Journal of Biological Chemistry, 1978, 253(6):1930-193.
Test principle
This ELISA kit uses the Sandwich-ELISA principle. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human TF. Samples (or Standards) and biotinylated detection antibody specific for Human TF are added to the micro ELISA plate wells. Human TF would combine with the specific antibody. Then Avidin-Horseradish Peroxidase (HRP) conjugate are added successively to each micro plate well and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human TF, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by the addition of stop solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 ± 2 nm. The OD value is proportional to the concentration of Human TF. You can calculate the concentration of Human TF in the samples by comparing the OD of the samples to the standard curve.
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Assay type |
Sandwich-ELISA |
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Format |
96T/48T |
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Assay time |
2.5h |
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Reactivity |
Human |
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Detection method |
Colormetric |
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Detection range |
0.14-100 ng/mL |
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Sensitivity |
0.13 ng/mL |
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Sample volume |
50μL |
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Sample type |
Serum, Plasma, Urine |
Specificity
This kit recognizes Human TF in samples. No significant cross-reactivity or interference between TF and analogues was observed.
Typical data
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.
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Concentration(ng/mL) |
100 |
33.33 |
11.11 |
3.70 |
1.23 |
0.41 |
0.14 |
0 |
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OD |
2.647 |
1.937 |
1.181 |
0.595 |
0.297 |
0.182 |
0.123 |
0.069 |
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Corrected OD |
2.578 |
1.868 |
1.112 |
0.526 |
0.228 |
0.113 |
0.054 |
- |
Sample values
Serum/Plasma/Urine–Samples from apparently healthy volunteers were evaluated for the presence of Human TF in this assay.
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Sample Type |
Source |
Range |
Dilution Factor |
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Serum（n=13） |
Healthy human |
2.58-4.22mg/mL |
2×105-4×106 |
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EDTA plasma（n=13） |
Healthy human |
2.13-4.42mg/mL |
2×105-4×106 |
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Urine（n=11） |
Healthy human |
51.45-1360.21ng/mL |
2-500 |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human TF were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human TF were tested on 3 different plates,20 replicates in each plate.
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Intra-assay Precision |
Inter-assay Precision |
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Sample |
1 |
2 |
3 |
1 |
2 |
3 |
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n |
20 |
20 |
20 |
20 |
20 |
20 |
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Mean(ng/mL) |
4.81 |
12.27 |
48.82 |
4.88 |
10.78 |
48.95 |
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Standard deviation |
0.24 |
0.6 |
2.4 |
0.26 |
0.49 |
2.36 |
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CV (%) |
4.99 |
4.89 |
4.92 |
5.33 |
4.55 |
4.82 |
Recovery
The recovery of Human TF spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
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Sample Type |
Range (%) |
Average Recovery (%) |
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Serum (n=13) |
87-94 |
91 |
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EDTA plasma (n=13) |
90-99 |
95 |
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Urine(n=11) |
95-107 |
100 |
Linearity
Samples were spiked with high concentrations of Human TF and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
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Serum (n=4) |
EDTA plasma (n=4) |
Urine(n=4) |
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1:3 |
Range (%) |
94-101 |
92-102 |
92-103 |
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Average (%) |
97 |
95 |
99 |
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|
1:9 |
Range (%) |
97-106 |
86-98 |
98-111 |
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Average (%) |
102 |
94 |
105 |
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1:27 |
Range (%) |
85-97 |
94-107 |
84-98 |
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Average (%) |
93 |
101 |
90 |
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|
1:81 |
Range (%) |
101-109 |
98-105 |
85-99 |
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Average (%) |
104 |
101 |
91 |
Kit components & Storage
An unopened kit can be stored at 2-8℃ for six months. After test, the unused wells and reagents should be stored according to the table below.
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Item |
Specifications |
Storage conditions after test |
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Micro ELISA Plate (Dismountable) |
96T: 8 wells ×12 strips 48T: 8 wells ×6 strips |
2-8℃, 1 month |
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Reference Standard |
96T: 2 vials 48T: 1 vial |
Discard unused reconstituted standard and dilutions |
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Reference Standard & Sample Diluent |
1 vial, 20 mL |
2-8℃ |
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Biotinylated Detection Ab Working Solution |
1 vial, 6 mL |
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HRP Conjugate Diluent |
1 vial, 14 mL |
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Concentrated Wash Buffer (25×) |
1 vial, 30 mL |
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Concentrated HRP Conjugate (100×) |
96T: 1 vial, 120 μL 48T: 1 vial, 60 μL |
2-8℃(Protect from light) |
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Substrate Reagent |
1 vial, 10 mL |
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Stop Solution |
1 vial, 10 mL |
2-8℃ |
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Plate Sealer |
5 pieces |
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Product Description |
1 copy |
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Certificate of Analysis |
1 copy |
Note: All reagent bottle caps must be tightened to prevent evaporation and microbial pollution.
The volume of reagents in partial shipments is a little more than the volume marked on the label, please use accurate measuring equipment instead of directly pouring into the vial(s).
Other supplies required
Microplate reader with 450 nm wavelength filter
High-precision transfer pipette, EP tubes and disposable pipette tips
37℃ Incubator
Deionized or distilled water
Absorbent paper
Loading slot for Wash Buffer
SUMMARY