Blood draw tubes: EDTA, Cell-free DNA BCT® (Streck)
Specifications
Technology: Silica membrane technology
Fragment size: ≥ 50 bp
NucleoSpin® DNA Plasma Midi
Processing: Vacuum
Sample size: Plasma (1–5 mL)
Elution volume: 200 μL (140 μL final eluate volume)
Processing time: 90 min/24 preps (EDTA plasma)
Principle / Procedure
NucleoSpin® DNA Plasma Midi is designed for the rapid, medium scale preparation of highly pure cell-free DNA from up to 5 mL blood plasma using vacuum filtration. The use of vacuum filtration technology eliminates tedious centrifugation steps and manual handling of columns during the process (e.g., discard of collected column flow through). Using the NucleoVac 96 Vacuum Manifold and the Starter Set Midi up to 24 samples can be processed in parallel. The Starter Set Midi consists of SBS format based column holders enabling the use of both manual vacuum manifolds, such as the NucleoVac 96 Vacuum Manifold as well as manifolds of liquid handling platforms. The set includes Column Holder Midi, Wash Plate Midi, and Elution Tube Holder Midi.
The purification protocol is based on the lysis of the plasma samples using Proteinase K incubation in the presence of a chaotropic salt containing buffer. Following the initial lysis step, a binding buffer is added to the samples and the lysate is loaded onto the binding columns. After loading and vacuum filtration driven binding of the DNA to the silica membrane, impurities are removed by several washing steps. cfDNA is eluted after a final drying step. Typically the processing time of 24 samples is approx. 90 min.
NucleoSpin® Plasma Midi procedure
Application data
Efficient isolation of cfDNA from 2 mL human EDTA plasma cfDNA was isolated from individual EDTA plasma samples with the NucleoSpin® DNA Plasma Midi kit and a vacuum-based kit from a competitor (Competitor Q). DNA yields were quantified by qPCR (Quantifiler® Human DNA Quantification Kit, ThermoFisher Scientific on an Applied Biosystems 7500 Real-Time PCR Systems ThermoFisher Scientific).
Optimized protocol for cfDNA isolation from Cell-free DNA BCT®(Streck) Blood was drawn from five independent test persons and preserved with Cell-free DNA BCT® and EDTA tubes. cfDNA was isolated with NucleoSpin® DNA Plasma Midi. cfDNA yield from EDTA and 'Streck tubes' are similar due to an optimized protocol for the isolation of cfDNA from 'Streck tubes'. DNA yields were quantified by qPCR (Quantifiler®Human DNA Quantification Kit, ThermoFisher Scientific on an Applied Biosystems 7500 Real-Time PCR System, ThermoFisher Scientific).
Blood draw tubes: EDTA, Cell-free DNA BCT® (Streck)
Specifications
Technology: Silica membrane technology
Fragment size: ≥ 50 bp
NucleoSpin® DNA Plasma Midi
Processing: Vacuum
Sample size: Plasma (1–5 mL)
Elution volume: 200 μL (140 μL final eluate volume)
Processing time: 90 min/24 preps (EDTA plasma)
Principle / Procedure
NucleoSpin® DNA Plasma Midi is designed for the rapid, medium scale preparation of highly pure cell-free DNA from up to 5 mL blood plasma using vacuum filtration. The use of vacuum filtration technology eliminates tedious centrifugation steps and manual handling of columns during the process (e.g., discard of collected column flow through). Using the NucleoVac 96 Vacuum Manifold and the Starter Set Midi up to 24 samples can be processed in parallel. The Starter Set Midi consists of SBS format based column holders enabling the use of both manual vacuum manifolds, such as the NucleoVac 96 Vacuum Manifold as well as manifolds of liquid handling platforms. The set includes Column Holder Midi, Wash Plate Midi, and Elution Tube Holder Midi.
The purification protocol is based on the lysis of the plasma samples using Proteinase K incubation in the presence of a chaotropic salt containing buffer. Following the initial lysis step, a binding buffer is added to the samples and the lysate is loaded onto the binding columns. After loading and vacuum filtration driven binding of the DNA to the silica membrane, impurities are removed by several washing steps. cfDNA is eluted after a final drying step. Typically the processing time of 24 samples is approx. 90 min.
NucleoSpin® Plasma Midi procedure
Application data
Efficient isolation of cfDNA from 2 mL human EDTA plasma cfDNA was isolated from individual EDTA plasma samples with the NucleoSpin® DNA Plasma Midi kit and a vacuum-based kit from a competitor (Competitor Q). DNA yields were quantified by qPCR (Quantifiler® Human DNA Quantification Kit, ThermoFisher Scientific on an Applied Biosystems 7500 Real-Time PCR Systems ThermoFisher Scientific).
Optimized protocol for cfDNA isolation from Cell-free DNA BCT®(Streck) Blood was drawn from five independent test persons and preserved with Cell-free DNA BCT® and EDTA tubes. cfDNA was isolated with NucleoSpin® DNA Plasma Midi. cfDNA yield from EDTA and 'Streck tubes' are similar due to an optimized protocol for the isolation of cfDNA from 'Streck tubes'. DNA yields were quantified by qPCR (Quantifiler®Human DNA Quantification Kit, ThermoFisher Scientific on an Applied Biosystems 7500 Real-Time PCR System, ThermoFisher Scientific).
