Air-Dryable™ Direct DNA and RNA/DNA qPCR Blood Master Mixes
Air-Dryable™ Direct DNA qPCR and RNA/DNA qPCR Blood mixes are glycerol-free mixes that contain optimized excipients compatible with air and oven drying and they are designed for the direct molecular quantitation of DNA and RNA from whole blood, serum, or plasma. Can also be used wet and/or with purified nucleic acid
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Meridian's Air-Dryable™ Direct DNA and RNA/DNA qPCR Blood Master Mixes
- Create ultra-sensitive multiplex direct blood qPCR tests using whole blood, serum or plasma.
- Simplify the development of quantitative PCR blood tests, by removing the nucleic acid extraction step and shortening the sample-to-result workflows.
- Validated for detection of blood pathogens and liquid biopsy applications.
- Can be used wet or air-dried for developing ambient-temperature stable assays.
Air-Dryable Direct DNA qPCR Blood, 4x, MDX092
Inhibitor tolerance to 20% whole blood, 20% plasma, compatible with all anticoagulants. Perfect for multiplexing and fast cycling conditions. Used in the detection of blood pathogens and liquid biopsy applications.
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Air-Dryable Direct RNA/DNA qPCR Blood, 4x, MDX121
Inhibitor tolerance to 10% whole blood, 10% plasma, compatible with all anticoagulants. Perfect for multiplexing and fast cycling conditions. Used in the detection of blood pathogens and liquid biopsy applications.
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Description
Blood is one of the most common specimens used for laboratory diagnostic testing and it is useful for evaluating the function of vital organs (kidneys, liver, thyroid, and heart) and for diagnosing diseases such as bacterial and viral infections, cancer, cardiovascular disease, and metabolic disorders such as diabetes. However, whole blood specimens, serum, and plasma contain a number of inherent PCR inhibitors including immunoglobulin G, hemoglobin, lactoferrin, and leukocyte DNA. In addition, PCR inhibitors can be found in the anticoagulants used to stabilize blood samples (e.g. EDTA, citrate, or heparin). Traditional methods have relied on removing these inhibitors by DNA or RNA extraction prior to testing, however, these methods are problematic, can cause sample loss, and are not 100% effective at removing all the inhibitors.
Air-Dryable™ DNA qPCR Blood is a 4x qPCR master mix for the development of DNA blood tests, enabling highly sensitive detection of target DNA from whole blood, plasma, and serum samples. Air-Dryable™ RNA/DNA qPCR Saliva is a 4x qPCR master mix and can be used for the development of both DNA and RNA blood tests from whole blood, plasma, and serum samples. These mixes have been formulated specifically to overcome the inhibitors found in blood samples – no further optimization is required aside from the addition of primers and probes. Furthermore, these mixes can be used in a wet format or oven dried to create ambient-temperature stable whole blood tests, plasma tests or serum tests.
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FAQs: qPCR Blood Test
What are the advantages of using blood over other sample types?
Blood contains abundant information about the health status of the organism, it can be used to evaluate organ function, detect proteins, nucleic acids, metabolites, infectious diseases and can be used for diagnosis of cancer (liquid biopsy), heart disease, diabetes. DNA and RNA Blood tests are a simple and easily accessible (low-invasive) method to assess and monitor health.
What qPCR inhibitors are found in whole blood?
Immunoglobulin G is the major inhibitor of qPCR in blood, as it can bind to single-stranded genomic DNA, leading to increased Ct values in a quantitative PCR blood test. The other two main inhibitors in blood are hemoglobin and lactoferrin that affect the DNA polymerase activity and thus lower the amplification efficiency. Hemoglobin and hematin can also cause some fluorescence quenching, particularly for FAM, although this should not affect the Ct value. In addition, inhibitors can be found in the anticoagulants used to stabilize blood samples (e.g. EDTA, citrate, or heparin).
What is the maximum volume of whole blood you can use in PCR blood Tests?
We have used up to 20% whole blood with our mix, higher concentrations cause to solution to become viscous and difficult to work with.
Do you use a mutant polymerase to overcome inhibitors?
We use our own proprietary Taq polymerase and a highly optimized buffer system to overcome inhibition.
After drying for how long will the mix stay stable to use? Or do we need to use it immediately after drying?
It is stable for up to 2 years at ambient temperature if correctly stored in sealed pouches.
Is it possible to detect cell free RNA/DNA from blood serum using these kits?
Yes, the Air-Dryable™ qPCR Blood kits can be used to detect both cfDNA and cfRNA, making it ideal for liquid biopsy.
Is a pre-treatment needed if you want to detect the DNA of gram-positive bacteria?
Yes, it is possible that you may need to pre-treat the blood in order to release the DNA from the gram-positive bacteria. In some cases a heat shock is sufficient, this will depend on the bacteria being examined.
What is the performance difference for DNA when using the DNA-specific blood mix versus the DNA/RNA blood mix?
We do not see a performance difference between the two mixes, which is why we call the Air-Dryable Direct RT-qPCR Blood mix – Air-Dryable™ Direct RNA/DNA qPCR Blood mix, as it can be used for both.
Can the blood with or without anticoagulant be store before the qPCR? How long? At what temperature?
It can be stored frozen or following the instructions from the storage buffer manufacturer.
Do the new master mixes need any changes in PCR protocol or are they compatible with the current setup?
They are compatible with standard cycling conditions and can also be used with very fast cycling conditions, however some optimization may be required for the amount of blood that is needed for qPCR blood tests.
Do I need to elute dried blood spots, or can they be used directly?
You should elute in water, or TE buffer.
Are reaction conditions the same for air-dried and liquid mixes?
Yes, the Air-Dryable™ Direct DNA qPCR Blood and Air-Dryable™ Direct RNA/DNA qPCR Blood can be used as a liquid mix or air/oven dried and stored for up to 24 months without needing to change the reaction conditions and air drying/ambient temperature storage will not affect the sensitivity of the test.
What type of pre-treatment is compatible with the inhibitor tolerant blood mixes (detergents, preheating mix enzyme/plasma 95°C 5 min in the PCR instrument)?
We advise not using Triton; however, heat treatment is ok.
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