Solution|Hangzhou Allsheng Instruments

cfDNA Extraction

Circulating free DNA (cfDNA) is a kind of DNA that is free outside the cell and cell-free, and widely exists in plasma, serum, urine, feces and cerebrospinal fluid. Most of the research applications mainly include cell free fetal DNA (cffDNA) and cell free tumor DNA (ctDNA), which are found in pregnant women and tumor patients respectively. Studies have shown that the amount of cfdna in blood increases under pathological conditions such as autoimmune diseases and cancer.

The average concentration of cfDNA in the plasma of healthy individuals ranged from 0 to 100 ng/mL, with an average concentration of 30 ng/mL, while the plasma cfDNA concentration of cancer patients was significantly increased and the concentration was often positively correlated with the stage of the cancer and the size of the tumor. Moreover, ctDNA has the same gene mutation type in tumor tissue and cell DNA, so the tumor cfDNA gene can be used as a new generation marker for non-invasive tumor detection. In addition, the change of ctDNA concentration during postoperative follow-up is closely related to the postoperative effect. A sharp decrease in ctDNA concentration after treatment means a better prognosis, and an increase in the concentration usually indicates the occurrence of drug resistance mutations or treatment failure. However, there are still certain challenges in using cfDNA to detect genes at this stage. First, as mentioned above, the content of cfDNA in blood is extremely low, so it is often necessary to extract and enrich from a few milliliters of peripheral blood. Second, the overall manual operation time is relatively long.

In order to improve the effect and efficiency of cfDNA extraction, Allsheng launched a full-process solution from sample extraction, concentration detection and automated library construction. Allsheng® magnetic bead method cell-free DNA (cfDNA) extraction kit is equipped with a variety of automated extractors, which can achieve max 4 mL samples automated extraction of cfDNA.

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cfDNA Extraction

cfDNA Extraction Kit

Instrument Features

· It can process samples from 200μL to 5mL to meet the extraction needs of different volumes samples
· Easy to operate, can be used for both manual and automated extraction
· Mature scheme, stable yield, no obvious fragment loss

Experimental Data

Nucleic Acid Extraction

Instrument Features

· The whole process automation design can realize reagent transfer, sample transfer, nucleic acid extraction and PCR set-up
· Large sample volume processing, maximum processing volume 10 mL (maximum sample volume 5 mL)
· Fast and accurate liquid transfer, dual-channel pipetting design, 1×50-5000μL, 1×1-200μL
· Easy-to-learn operating software, graphical user interface, and program editing without complicated operations
· Specially designed kits can be packaged separately and can extract and purify nucleic acid from a single sample
· Open software design can be adapted to various cfDNA extraction kits

Experimental Data

Concentration Measurement

Instrument Features

· Fluo-200/800 fluorometer can detect 1/8 samples in one run
· Accurate quantification, only 1-20μL can be used to obtain accurate sample concentration
· High sensitivity, the lowest detection limit can reach 1pg/μL (dsDNA)
· Open design, suitable for all kinds of fluorometer detection reagents

Experimental Data

cfDNA Extraction Kit
Nucleic Acid Extraction
Concentration Measurement

cfDNA Extraction Kit

Bioanalyzers characterize the distribution of each fragment of the extracted product

Fragment analysis was performed on 1 ul of the original DNA ladder sample and 1 ul of the recovered DNA sample at the same time. The results showed that the fragment distribution of the two was highly consistent, and the recovery efficiency was over 90%.

Plasma extraction results of tumor patients with different kits

Allsheng®, imported kit T and domestic kit A were used to extract 2×1 ml tumor patient plasma samples respectively, and finally the magnetic beads were eluted in 50 ul eluent, and the concentration was tested by Fluo-100 fluorometer.
Nucleic Acid Extraction
Concentration Measurement

The results of plasma extraction from tumor patients with different volumes

The plasma of 0.5 ml, 1 ml and 4 ml of tumor patients was extracted by manual method and automated instrument respectively, and the elution volume was 50 ul. The results showed that there was no significant difference in the yields between the two extraction methods under the three volumes (a), and the fragment size distributions were roughly the same (b, c, d).