Automated isolation of cell-free tumor or fetal DNA
from 3 - 5 ml plasma
In the event of apoptosis, necrosis or break-down of pathogens, or cells, such as white blood cells cfDNA is floating through the blood. As the average size of cfDNA is 170 bp (50 – 200 bp) and only 1 – 50 ng of cfNA/ml of plasma is present, it is quite difficult to separate this hay in the haystack. Nevertheless the detection of cfDNA in the blood is an elegant non-invasive method to detect or monitor tumors or predict chromosomal disorders without surgery. PerkinElmer chemagen has a new method to isolate cfDNA form 3-5 ml plasma within 80 minutes with the chemagic 360 instrument.
Kit for chemagic™ automation
|Catalog no.||Kit name||Instrument||Input||Typical yield||Format||No. of preps|
|CMG-1304||chemagic™ cfDNA 5k Kit H24||chemagic™ 360||3 - 5 ml||1-30 ng/ml plasma (qPCR)||24 XL plate||240|
The kit is designed for the use with human plasma samples derived from EDTA, citrate or Streck Cellfree DNA BCT® tubes.
Fresh and frozen plasma can be used.
With the chemagic cfDNA 5k Kit H24 circulating cell-free DNA (cfDNA) can be isolated from 3 - 5 mL of plasma obtained from human whole blood samples. All the reagents needed for the isolation of cfDNA are included in the kit, with the exception of nuclease-free water for dissolving Proteinase K. The kit components (reagents and plastic ware) provide sufficient material for 240 preparations. The kit is designed to be used with the chemagic 360 and integrated chemagic Dispenser. The product is intended for professional users such as technicians and physicians trained in molecular biology techniques.
The chemagic cfDNA 5k Kit H24 is based on chemagic Technology using M-PVA Magnetic Beads for the isolation of cfDNA. The cfDNA binds to paramagnetic beads which are magnetically separated from the sample material. During subsequent steps contaminants are removed and the purified cfDNA is transferred into an elution medium. The automated sample processing by the chemagic 360 excludes cross contamination and ensures safe handling of infectious sample material.
Plasma Preparation Protocol
Plasma Preparation Protocol
It is recommended to prepare plasma as fresh as possible (max. 5 days after blood draw). Longer storage of blood prior to plasma preparation may lead to poor separation results and higher background from genomic DNA.
A double centrifugation protocol during plasma preparation is recommended to minimize the potential for contamination of plasma with cells and genomic DNA. Transfer of any other blood components (buffy coat or red blood cells) should be avoided while separating the plasma fraction.
Please refer to the tube manufacturers specifications (max. centrifugation speed) for more information.
From 10 mL of whole blood approximately 4 - 5 mL plasma can be expected. Plasma volumes below 3 mL cannot be used for cfDNA extraction using this kit.
- Centrifuge whole blood collection tube at 2.000 x g for 20 minutes.
- Aspirate plasma carefully and at least 2 - 3 mm above the buffy coat layer, without disturbing the layer and transfer it into a new appropriate tube.
- Centrifuge the plasma sample at 3.300 x g for 30 minutes.
- Carefully transfer the supernatant to a fresh tube without disturbing the pellet, at least 2 - 3 mm above the pellet.
- For direct use storage of plasma sample at 2 - 8 °C is possible, for long-term storage -80 °C is recommended.
For research use only. Not for use in diagnostic procedures.