Specimen-specific mixes designed for direct qPCR and RT-qPCR from stool specimens. Lyo-Ready™ Direct DNA qPCR and RNA/DNA qPCR Stool mixes are 4x concentrated, glycerol-free mixes that contain excipients compatible with lyophilization and are designed for sensitive amplification of RNA or DNA from crudely processed stool specimens.
Lyo-Ready Direct Stool Mixes Specimen-specific mixes designed for direct qPCR and RT-qPCR from stool specimens
Lyo-Ready ™ Direct DNA qPCR and RNA/DNA qPCR Stool mixes are 4x concentrated, glycerol-free mixes that contain excipients compatible with lyophilization and are designed for sensitive amplification of RNA or DNA from crudely processed stool specimens. Fecal matter is a highly complex, heterogenous material that contains substances which can interfere with PCR such as food debris, desquamated epithelial cells and mucus, and PCR inhibitors including bile salts, polysaccharides, hematin and catabolic substances. Generally, stool samples must undergo substantial processing to extract the DNA or RNA prior to testing in order to avoid PCR inhibition. Lyo-Ready ™ Direct DNA qPCR and RNA/DNA qPCR Stool mixes are uniquely designed for direct qPCR/RT-qPCR analysis, and contain an optimized blend of additives to negate PCR inhibitor effects while maintaining the quality and integrity of nucleic acids found in patient samples. In addition, the mixes are glycerol-free and are suitable to use in a liquid format or they can be lyophilized to create ambient-temperature stable assays. As ready-to-use mixes, no further optimization is required aside from the addition of disease-specific primers and probes. Lyo-Ready ™ Direct DNA qPCR and RNA/DNA qPCR Stool mixes simplify the process of developing direct detection stool assays, and as the need for fast, non-invasive testing for gastrointestinal conditions increases, short turn-around times, high sensitivity and a long shelf-life become important distinguishing features.
Product Highlights
• Simple and easy to use • Suitable for direct detection of very low titers of RNA or DNA (including ctDNA) in singleplex or multiplex assays using stool samples • Mixes can be used as a liquid or lyophilized, reducing the cost and complexity of developing an ambient-temperature stable assay • Compatible with a range of lyophilization protocols
PRODUCT
CAT NO. VOLUME REACTIONS
5 mL 1,000 Rxns 50 mL 10,000 Rxns 5 mL 1,000 Rxns 50 mL 10,000 Rxns
Lyo-Ready ™ Direct DNA qPCR Stool
MDX142
Lyo-Ready ™ Direct RNA/DNA qPCR Stool
MDX143
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Sensitive Detection in Multiplex Assays in the Presence of Stool Inhibitors
Lyo-Ready™ Direct DNA qPCR Stool (MDX142)
Escherichia coli O157
Salmonella typhimurium
Campylobacter jejuni
Lyo-Ready ™ Direct DNA qPCR Stool | KAPA Probe Force | UltraPlex ™
E. coli O157 (Shiga-like/Verotoxin 2 gene), Salmonella typhimurium (16S-23S rRNA gene internal transcribed spacer) and Campylobacter jejuni (16S-23S rRNA gene internal transcribed spacer) were amplified in a triplex reaction using lyophilized Lyo-Ready ™ Direct DNA qPCR Stool ( red ) and universal mixes, KAPA Probe Force (Roche, green ) and UltraPlex ™ (QuantBio, grey ) in the presence of 6.6 mg/mL human stool. The results illustrate that Lyo-Ready ™ Direct DNA qPCR Stool is not affected by high concentrations of human stool, unlike the other mixes, allowing higher multiplexing capacity and greater reproducibility.
Lyo-Ready™ Direct RNA/DNA qPCR Stool (MDX143)
Rotavirus
Norovirus
Astrovirus
Campylobacter jejuni
Lyo-Ready ™ Direct RNA/DNA qPCR Stool | TaqPath ™ | UltraPlex ™
Three RNA targets (Rotavirus, Norovirus and Astrovirus) and one DNA target ( Campylobacter jejuni ) were amplified in a quadruplex reaction using lyophilized Lyo-Ready ™ Direct RNA/DNA qPCR Stool ( red ) and universal mixes, TaqPath ™ (Thermo, black ) and UltraPlex ™ (QuantBio, grey ) in the presence of 1.5 mg/mL bile salts. The results illustrate that unlike TaqPath ™ and Ultraplex ™ which are universal mixes, Lyo-Ready ™ Direct RNA/DNA qPCR Stool is less affected by high concentrations of bile salts, allowing higher multiplexing capacity and greater reproducibility.
Ideal for Liquid Format or Lyophilized Assays – No Impact on Reaction Efficiency
A) Lyo-Ready ™ Direct DNA qPCR Stool (MDX142)
B) Lyo-Ready ™ Direct RNA/DNA qPCR Stool (MDX143)
10,000 copies 1,000 copies
10,000 copies 1,000 copies
100 copies 10 copies
100 copies 10 copies
lyophilized | liquid
Liquid ( blue ) and lyophilized ( red ) formats of A) Lyo-Ready ™ Direct DNA qPCR Stool (MDX142) was used in a 10-fold serial dilution of DNA (10,000, 1,000, 100, and 10 copies respectively) in presence of 20% artificial stool and B) Lyo-Ready ™ Direct RNA/DNA qPCR Stool (MDX143) was used in a 10-fold serial dilution of RNA (10,000, 1,000, 100 and 10 copies respectively) in presence of 5% artificial stool. The results illustrate that the lyophilized mixes retain the ability to efficiently amplify to the same level as the liquid mixes with the same level of sensitivity and reproducibility.
High Performance with Both Standard and Fast Cycling Conditions Provides Greater Assay Flexibility and Reduced Run Times
Lyo-Ready™ Direct DNA qPCR Stool (MDX142)
Standard cycling
Fast cycling
Two mammalian DNA targets ( red ) were amplified using standard qPCR cycling conditions and the fastest cycling conditions available on a Quantstudio ™ 7 Flex Real-Time PCR System in the presence of 6.6 mg/mL human stool. The results illustrate the higher performance and end-fluorescence of Lyo-Ready ™ Direct DNA qPCR Stool (MDX142) in both standard (3 min @ 95ºC, 40 cycles of 10 sec @ 95ºC and 30 sec @ 60ºC) and fast (3 min @ 95ºC, 40 cycles of 1 sec @ 95ºC and 1 sec @ 60ºC) qPCR cycling conditions, without compromising the sensitivity and reproducibility of an assay.
www.meridianbioscience.com/lifescience
Lyo-Ready™ Direct RNA/DNA qPCR Stool (MDX143)
Standard cycling
Fast cycling
Two mammalian RNA targets ( red ) were amplified using standard qPCR cycling conditions and the fastest cycling conditions available on a Quantstudio ™ 7 Flex Real-Time PCR System in the presence of 1.5 mg/mL human stool. The results illustrate the higher performance and end-fluorescence of Lyo-Ready ™ Direct RNA/DNA qPCR Stool (MDX143) in both standard (10 min reverse transcription step followed by 40 cycles of 10 sec @ 95ºC and 30 sec @ 60ºC) and fast (5 min reverse transcription step followed by 40 cycles of 1 sec @ 95ºC and 1 sec @ 60ºC) qPCR cycling conditions, without compromising the sensitivity or reproducibility of the assay.
Stable Shelf-life for 12+ Months in Lyophilized Format
A)
B)
Lyophilized | Liquid
Lyo-Ready Direct DNA qPCR Stool (MDX142) and Lyo-Ready Direct RNA/DNA qPCR Stool (MDX143) were lyophilized, and their stability was tested in an accelerated stability study. The lyophilized mixes ( red ) were incubated at 37ºC for 4 weeks and tested against freshly prepared liquid mixes ( blue ) in A) a qPCR assay with 20% artificial stool or B) an RT-qPCR assay with 5% artificial stool. The lyophilized mixes retained the same activity as the liquid mixes (<0.5 Δ Ct) over time. Results indicate that the lyophilized mixes have a projected stability of at least 12 months at ambient temperature.
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Email: info@meridianlifescience.com Orders: orders@meridianlifescience.com www.meridianbioscience.com/lifescience
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