Ordering (12)
We occasionally makes updates to our catalog, resulting in changes to or discontinuation of previously offered products. To know specifically why a product is unavailable for re-order through your order history, and to discuss options, please contact us.
We take pride in ensuring that all products customers receive are developed and manufactured to the highest standards.
The MMolecular products are made according to international standards of DIN EN ISO 9001.
MMolecular accepts purchase orders (PO), direct bank transfer and all major credit cards (Visa, MasterCard, and American Express).
If you cannot find the assay you are looking for, MMolecular offers a service for assay development. All of our assays are also available in a multiplex configuration. Let us know your specific multiplex needs and we will get back to you.
All payments are due within 30 days of the invoice date. We accept purchase orders (PO), direct bank transfer and all major credit cards (Visa, MasterCard, and American Express).
If the order has not been processed, we can cancel the order, allowing you to reorder with the additional products you require. We cannot add products to Purchase Orders (PO).
MMolecular will email you a confirmation when your order has been shipped. This shipping confirmation will include the tracking number of the package, which you can use to monitor the status of your shipment. This shipping confirmation email is usually sent to the same email address at which you receive order confirmations.
An email order confirmation is sent to the email address on the account shortly after an order is received and processed. If you do not receive this email, please contact us.
To review your order history, click on MY ACCOUNT and sign in. You will then be able to consult your order history.
MMolecular accepts purchase orders (PO), direct bank transfer and all major credit cards (Visa, MasterCard, and American Express).
We cannot accept orders through email. This is to ensure that addresses, shipping options, payment details and ordered products are correctly entered the way you want. We offer 3 convenient ways to order: from our online catalogue, through the QUICK ORDER FORM and reorder from your order history.
The ability to modify an order depends on the type of order and the change you want to make. Any requests made before processing of your order has begun can usually be accommodated.
RT-qPCR (17)
Check your real-time PCR instrument to make sure it is set at the correct wavelength to detect the FAM probe and the cycling parameters are set properly.
Try to detect the amplicon on an electrophoresis gel.
We recommend that you repeat a failed experiment once to make sure it was not due to a simple mistake in the first round.
For accurate analysis of qPCR results, each experiment needs to be set up with multiple controls.
While the exogenous positive controls that we offer are useful, they are intended to create calibration curves for quantitative analysis. You may not need quantified results, if only confirming a positive or negative reaction. This allows for a more cost efficient kit.
To be fully confident, it might make more sense to incorporate a know positive sample as a positive control. This will enable to verify the entire sample processing chain and avoid any mistakes.
Under proper conditions, fluorescein-based dyes (e.g., the standard 6-FAM dye used for the probes in our kits) have excellent signal intensity. However, they are susceptible to a relatively high rate of photobleaching and are sensitive to acidic environments (ideal pH is 7.2 or higher), which can diminish the signal over time.
To minimize photobleaching, we recommend using a maximally sensitive detection system with the lowest possible intensity of excitation.
We aim for a final concentration of 150-250 nM of each probe in the reaction.
We aim for a final concentration of 500 nM of each primer in the reaction.
Ensuring that qPCR is quantitative, accurate, and reproducible requires thoughtful assay design, setup, and optimization. During design, each of the MMolecular RT-qPCR assays follows stringent criteria to ensure optimal performance.
Each MMolecular assay meets the following criteria:
-PCR efficiency 2.0 +/- 0.2 (equals 100 +/- 10 %).
-Cq of highest cDNA concentration <= 34.
-Linear dynamic range of at least 3 logs.
-High amplification specificity, with no side products in gel analysis.
-Sigmoidal amplification curve.
-Fluorescence intensity of amplification curves between 5 and 50 fluorescence units.
In the direct one-step method, there is very little sample processing. The extraction process generally is done in the PCR tube. Eg. reactions from blood samples can be run directly. The reverse transcriptase is also included in the same tube as the PCR. This saves time and reduces the possibility of sample contamination. A two step reaction will have better sensitivity.
The one-step method requires RNA isolation prior to running the PCR reaction. The reverse transcriptase and PCR reactions are run in the same tube. This saves time and reduces the possibility of sample contamination. A two step reaction will have better sensitivity.
The two-step method requires RNA isolation, next a reverse transcriptase reaction and then the actual PCR reaction can be done. Two-step RT-PCR is highly sensitive, potentially more efficient, and allows for stocking of the cDNA for future use. However, this method is more time consuming and can require more optimization.
Our protocols assume that you will be using a 20µL reaction. We recommend that you use 20µL as a general staring point. A larger volume may have better sensitivity than a smaller volume.