F-E3 estriol (free) ELISA kit
US DRG imported, article number: EIA-1612
US DRG China exclusive agent "Shenzhen Kerunda Biological Engineering Co., Ltd." dedicated to serve you
1 , the scope of application
DRG's estriolase-free kit can be used for in vitro detection of serum free estriol in pregnant women during pregnancy.
2 , the experimental description
Estriol (E3) is the major estrogen produced by the placenta during pregnancy. Free E3 enters the mother's blood circulation through the placenta, where it quickly degrades into glucuronide and sulfate derivatives for excretion. The half-life of estriol in the mother's blood is only 20-30 minutes, so the recent fetal status can be easily and quickly evaluated. Plasma estriol levels steadily increased and rose fastest in the third trimester (28-40 weeks). A sudden decrease in the production of placental estriol will result in a rapid decrease in free estriol in the mother's serum. Determination of free estriol has many possible advantages over the determination of serum total E3 or urinary E3, free estriol levels and maternal liver and kidney. It is not related to function and is not affected by certain antibodies, so free estriol more accurately reflects the outcome of pregnant women with diabetes. Because free estriol does not undergo any hydrolysis, we can get experimental results faster. DRG's E3 enzyme-free kit detects free E3 in human serum in a non-isotopic manner. This method utilizes a highly specific E3 antibody and enzyme-linked analyte. Finally, the color product was detected by a microplate reader. This kit has a long shelf life, no radioisotope, and is certified, so it is widely used in laboratories of all sizes.
3 , the principle of experiment
DRG's estriol (free) enzyme-free kit is a solid-phase enzyme-free adsorption assay (ELISA) based on the principles of competition. In this experiment, estriol was labeled with horseradish peroxidase. Serum containing estriol was mixed with horseradish peroxidase-labeled estriol and added to the microwells of the coated plates. The antibody to estriol has been pre-coated on the microwell of the coated plate. After one hour of incubation, unbound horseradish peroxidase-labeled estriol and sample estriol were washed away with washings. The substrate solution was further added to the micropores, and after the incubation, a stop solution capable of producing a colored product was added, and then the OD value was read on a microplate reader.
3 , kit components
Note: Store in 2-8 ° C environment.
1) Rabbit anti-estrogen antibody coated coated plate, 12 × 8 well, coated with estriol antibody in microwell.
2) Serum triol serum standard, a total of 6 (standard 0-5), 1ml / support, (0; 0.3; 1.2; 4; 15; 40ng / ml), each with a person containing estriol Serum, salt buffer and preservatives. Note: Treat it as a potential infectious agent for hepatitis B virus and HIV. The raw materials of the standards were tested with recognized reagents and found to be non-reactive with hepatitis B virus surface antigen and HIV. However, there is currently no way to ensure that products derived from humans do not contain the above viruses.
3) Estradiol enzyme conjugate, 14 m, containing horseradish peroxidase-labeled estriol.
4) TMB substrate solution, 14 ml.
5) Stop solution, 14ml, H2SO4.
6) Concentrated washing solution (40X), 30ml.
Note: The kit may contain materials that require special handling like human serum. Before disposing of the remaining materials, pay attention to local regulations regarding the handling of medical waste to ensure that the treatment methods and results meet the requirements of local laws and regulations.
4 , the virtual material of the experiment (but the kit does not provide)
1) Standard pipette
2) Microplate reader that can read at 450nm
3) Absorbent paper
5 , sample collection and processing
1) Treat all blood and serum as potential sources of hepatitis B virus and HIV infection.
2) Serum is required during the detection of E3.
3) Separate the serum immediately after collecting the blood, and cover it with a lid when storing.
4) The sample can be stored for 4 days at 2-8 °C. If it is to be stored for more than 4 days, it must be stored frozen.
5) The sample should be protected from repeated freezing and thawing.
6) The frozen serum must be fully thawed and mixed well before the start of the experiment.
7) Do not use serum with high levels of fat. (At the beginning of the experiment, you must try to detect the lipid content of the serum).
8) Samples containing a certain amount of lipemia, hemolysis, and xanthine blood generally do not affect the results of the experiment.
6 , preparation of washing liquid
The concentrated washing liquid was diluted with 900 ml of distilled water, and stored in an environment of 2-8 ° C, and was stable during the effective period.
7 , experimental steps
1) Add 10 μl of estriol standard to the corresponding microwells.
2) Add 10 μl of the control and patient serum to the corresponding microwells.
3) Add 100 μl of enzyme-labeled estriol to each well.
4) Mix well for 10 seconds.
5) Incubate for 60 minutes at room temperature.
6) Discard the reaction solution in the well, wash the plate 4 times with 400 μL of washing solution per well, and pat dry on absorbent paper.
7) Add 100 μl of TMB substrate solution to each well.
8) Incubate for 30 minutes at room temperature.
9) Add 100 μl of Stop Solution to each well.
10) Read the OD value at 450 ± 10 nm.
8 , experimental considerations
1) When adding reagents, ensure that the order of loading of each well is the same, which ensures that all wells have the same incubation time. Each loading step should be uninterrupted.
2) To prevent liquid from remaining on the pore walls, add the reagent directly to the bottom of the well.
3) In order to obtain the desired experimental results, our test standards and samples are double tested.
4) The pipettes used in the experiments, including multichannel or automatic pipettes, must have a CV (coefficient of variation) of less than 1%.
9 , results
9.1 manual calculation
1) On the semi-logarithmic coordinate paper, the OD value of the E3 standard is the Y-axis, and the concentration is the X-axis as a standard curve.
2) Draw a best curve between points
3) On the standard curve, plot the corresponding concentration of the OD value of the control and the unknown serum sample, and record the result.
DRG's MAT3000 microplate reader and corresponding linear regression curves can be used to directly read OD values ​​using computer-assisted four-parameter and three-regression curves.
10 , operational limitations
The user of the kit must read and fully understand the instructions before the start of the experiment. Reliable experimental results can be obtained by strictly following the experimental procedures. Care must be taken when handling, storing, and adding reagents, which is important for the accuracy and precision of the results. Improper handling of patient samples can lead to erroneous experimental results. Before the experiment begins, mix the frozen and thawed samples thoroughly. Do not use stale and mishandled serum samples. Degradation of the sample and repeated freezing and thawing can lead to erroneous experimental results. Patient samples should be tested as soon as possible. Samples with severe hemolysis, lipemia, or jaundice will result in lower precision and precision. The experimental results obtained with this sample should be handled with care. Experiments This should use better quality samples to determine the results.
11, experimental quality control instructions
1) Do not mix or exchange different kits or different batches of reagents.
2) Do not use expired reagents. (The expiration date is printed on the reagent bottle)
3) Each laboratory should use quality control to determine the precision reading and accuracy standards of its laboratory in the normal range, low level range and high level.
4) In order to ensure the reliability and continuity of the results of different batches of kits, the drawing method and statistical methods should be continuously updated.
12 , clinical significance
Detection of human body fluid E3 is often used to monitor the growth of the fetus, especially for high-risk women. The concentration of plasma E3 will gradually increase during the 20th week of pregnancy and will increase more rapidly during the second trimester. Because the normal and abnormal range of serum E3 is quite broad and unstable, the detection of E3 alone has no significant clinical significance. In order to determine the trend of individual E3, we should always monitor the patient's condition. Serum E3 levels in late pregnancy continue to be low or abrupt and continue to decrease, indicating a high likelihood of fetal distress or uterine stillbirth. If this happens, the corresponding method should be used to detect the condition of the fetus. Serum free E3 levels Invar results in other clinical methods or diagnostic methods (amniocentesis and ultrasound) to illustrate its clinical significance. Abnormal levels of E3 may also be detected in the serum of patients with antibiotics or corticosteroids and patients with impaired maternal liver function.
13 , expected value
Gestational week | Reference range (ng/ml) | Gestational week | Reference range (ng/ml) | Twin pregnancy (ng/ml |
12 | 0.3-1.0 | 22-23 | 2.7-16 | 3-18 |
13 | 0.3-11 | 24-25 | 2.9-17 | 3-20 |
14 | 1.4-1.6 | 26-27 | 3.0-18 | 4-21 |
15 | 1.0-4.4 | 28-29 | 3.2-20 | 4-22 |
16 | 1.4-6.5 | 30-31 | 3.6-22 | 5-25 |
17 | 1.5-6.6 | 32-33 | 4.6-23 | 6-39 |
18 | 1.6-8.5 | 34-35 | 5.1-25 | 7-39 |
19 | 1.9-11 | 36-37 | 7.2-29 | 9-38 |
20 | 2.1-13 | 38-39 | 7.8-37 | 13-40 |
twenty one | 2.6-14 | 40-42 | 8.0-39 | -- -- |
Exclusive distributor in China: Shenzhen Kerunda Bioengineering Co., Ltd.
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