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Anti-Mullerian Hormone (AMH) ELISA Kit
|Synonyms||MIF, MIS, AMH, amh|
|1 reference available|
|Certificates||ISO 9001:2008, ISO 13485:2003|
|Price||729.47 $ Plus shipping costs $45.00|
|Availability||Will be delivered in 9 to 11 Business Days|
|Method type||Competition ELISA|
|Cross-Reactivity (Details)||No significant cross-reactivity or interference between human AMH and analogues was observed. Note: Limited by current skills and knowledge, it is impossible for us to complete the cross- reactivity detection between human AMH and all the analogues, therefore, cross reaction may still exist.|
|Sample Type||Serum, Plasma, Biological Fluids, Tissue Homogenate, Cell Lysate, Cell Culture Supernatant|
|Plate||Pre-coated 12 × 8 strip|
|Specificity||This assay has high sensitivity and excellent specificity for detection of human AMH.|
|Sensitivity||The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest protein concentration that could be differentiated from zero. It was determined by subtracting two standard deviations to the mean optical density value of twenty zero standard replicates and calculating the corresponding concentration.|
|Minimum Detection Limit||15.6 pg/mL|
|Detection Range||37.0-3,000 pg/mL|
Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level human AMH were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level human AMH were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV < 10%
Inter-Assay: CV < 12%
The standard curve concentrations used for the ELISA’s were 3,000pg/mL, 1,000pg/mL, 333.3pg/mL, 111.1pg/mL, 37.0pg/mL.The stability of ELISA 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.
|Principle||The kit is a competitive inhibition enzyme immunoassay technique for the in vitro quantitative measurement of AMH in human serum, plasma, tissue homogenates, cell lysates, cell culture supernates and other biological fluids.|
This assay employs the competitive inhibition enzyme immunoassay technique. A monoclonal antibody specific for human AMH has been pre-coated onto a microplate. A competitive inhibition reaction is launched between biotin labeled human AMH and unlabeled human AMH (Standards or samples) with the pre-coated antibody specific for human AMH. After incubation the unbound conjugate is washed off. Next, avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. The amount of bound HRP conjugate is reverse proportional to the concentration of AMH in the sample. After addition of the substrate solution, the intensity of color developed is reverse proportional to the concentration of AMH in the sample.
Serum: Use a serum separator tube and allow samples to clot for two hours at room temperature or overnight at 4°C before centrifugation for 20 minutes at approximately 1000×g. Assay freshly prepared serum immediately or store samples in aliquot at -20°C or -80°C for later use. Avoid repeated freeze/thaw cycles.
Plasma: Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples for 15 minutes at 1000×g at 2 - 8°C within 30 minutes of collection. Remove plasma and assay immediately or store samples in aliquot at -20°C or -80°C for later use. Avoid repeated freeze/thaw cycles.
Tissue homogenates: The preparation of tissue homogenates will vary depending upon tissue type. For this assay, tissues were rinsed in ice-cold PBS(0.02mol/L,pH 7.0-7.2) to remove excess blood thoroughly and weighed before homogenization. Minced the tissues to small pieces and homogenized them in 5-10 mL of PBS with a glass homogenizer on ice(Micro Tissue Grinders woks, too). The resulting suspension was sonicated with an ultrasonic cell disrupter or subjected to two freeze-thaw cycles to further break the cell membranes. After that, the homogenates were centrifugated for 5 minutes at 5000×g. Remove the supernate and assay immediately or aliquot and store at ≤ -20°C.
Cell Lysates: Cells must be lysed before assaying according to the following directions. 1. Adherent cells should be detached with trypsin and then collected by centrifugation (suspension cells can be collected by centrifugation directly). 2. Wash cells three times in cold PBS. 3. Resuspend cells in PBS (1×) and the cells was subject to ultrasonication for 4 times (or Freeze cells at ≤ -20°C. Thaw cells with gentle mixing. Repeat the freeze/thaw cycle for 3 times.) 4. Centrifuge at 1500×g for 10 minutes at 2 - 8°C to remove cellular debris. Cell culture supernates and other biological fluids - Centrifuge samples for 20 minutes at 1000×g. Remove particulates and assay immediately or store samples in aliquot at -20°C or -80°C for later use. Avoid repeated freeze/thaw cycles.
|Calculation of Results||This assay employs the competitive inhibition enzyme immunoassay technique, so there is an inverse correlation between AMH concentration in the sample and the assay signal intensity. Average the duplicate readings for each standard, control, and samples. Create a standard curve on log-log or semi-log graph paper, with the log of AMH concentration on the y-axis and absorbance on the x-axis. Draw the best fit straight line through the standard points and it can be determined by regression analysis. Using some plot software, for instance, curve expert 1.30, is also recommended. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.In order to make the calculation easier, we plot the O.D. value of the standard (X-axis) against the log of concentration of the standard (Y-axis), although concentration is the independent variable and O.D. value is the dependent variable. The O.D. values of the standard curve may vary according to the conditions of assay performance (e.g. operator, pipetting technique, washing technique or temperature effects). Typical standard curve below is provided for reference only.|
Limited by the current condition and scientific technology, we cant completely conduct the comprehensive identification and analysis on the raw material provided by suppliers. So there might be some qualitative and technical risks to use the kit.
The final experimental results will be closely related to validity of the products, operation skills of the end users and the experimental environments. Please make sure that sufficient samples are available.
Kits from different batches may be a little different in detection range, sensitivity and color developing time.
Do not mix or substitute reagents from one kit lot to another. Use only the reagents supplied by manufacturer.
Protect all reagents from strong light during storage and incubation. All the bottle caps of reagents should be covered tightly to prevent the evaporation and contamination of microorganism.
There may be some foggy substance in the wells when the plate is opened at the first time. It will not have any effect on the final assay results. Do not remove microtiter plate from the storage bag until needed.
Wrong operations during the reagents preparation and loading, as well as incorrect parameter setting for the plate reader may lead to incorrect results. A microplate plate reader with a bandwidth of 10nm or less and an optical density range of 0-3 O.D. or greater at 450 ± 10nm wavelength is acceptable for use in absorbance measurement. Please read the instruction carefully and adjust the instrument prior to the experiment.
Even the same operator might get different results in two separate experiments. In order to get better reproducible results, the operation of every step in the assay should be controlled. Furthermore, a preliminary experiment before assay for each batch is recommended.
Each kit has been strictly passed Q.C test. However, results from end users might be inconsistent with our in-house data due to some unexpected transportation conditions or different lab equipments. Intra-assay variance among kits from different batches might arise from above factors, too.
Kits from different manufacturers for the same item might produce different results, since we havent compared our products with other manufacturers.
|Handling Advice||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.|
|Precaution of Use||The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.|
|Material not included||
|Expiry Date||The expiry date is stated on the label.|
|Restrictions||For Research Use only|
Lin, Wang, Weng et al.: "Influence of gonadotropin-releasing hormone agonist on the effect of chemotherapy upon ovarian cancer and the prevention of chemotherapy-induced ovarian damage: an experimental study with nu/nu athymic mice." in: Journal of Zhejiang University. Science. B, Vol. 13, Issue 11, pp. 894-903, 2012 (PubMed).