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2023 - A look back | High Throughput cfDNA Analysis

Katharina Esch

UpToDate and LexiDrug Senior Account Manager Nordics I Wolters Kluwer Health

发布日期: 2023年12月6日

The year is slowly coming to an end. And what a year it has been. The big change happened in May when it was announced that we are now officially a new business, we are Revvity.

This newsletter shall therefore recap the year 2023 and the last 10 issues of this year`s newsletter but also briefly mention one very exciting high throughput cfDNA analysis you can use our LabChip GX instrument for. Maybe something you would like to test in the new year as we already see a lot of cfDNA work going on in the Nordics.

LabChip? GX Touch? cfDNA Assay for High Throughput Analysis of Cell-free DNA

Unlike many other assays which use a higher base molecular weight upper marker for DNA quantitation, the cfDNA assay uses a 50 base pair DNA as an internal standard that is similar in size to cfDNA smears.

This analysis method corrects sample injection bias enabling improved quantitation accuracy. Faster analysis and higher assay sensitivity is achieved by using optimized gel matrix conditions.

The LabChip? cfDNA Assay is capable of detecting cfDNA samples at as low as 25 pg/μL concentration (S/N>3) and has a linear range of 50-1000 pg/μL.

Process time for 96 samples is 90minutes.

cfDNA samples were extracted from pooled plasma reformatted from whole blood tubes by Janus Blood iQ using different extraction kits [chemagic? cfDNA 1.5k Kit (PerkinElmer, Part # CMG-1396), MagMAX? Cell-Free DNA Isolation Kit (Thermo Fisher Scientific, Part # A29319), chemagic? cfDNA 5k Kit (PerkinElmer, Part # CMG-134), QIAamp? Circulating Nucleic Acid Kit (Qiagen, Part # 55114)] according to manufacturer-recommended protocols.

Comparison of Assay Specifications of LabChip cfDNA Assay and Competitor A Assay (closest in performance available in the market)

Analysis Time, Sensitivity, & Linear Range

Each sample run is completed in 40 s with LabChip cfDNA assay as seen in the electropherogram in Figure 3A, (2.5 times faster compared to the competitor method); a total of 96 samples can be analyzed in 1 hour and 26 minutes (instrument warm-up time included) in one chip preparation in high-throughput mode. Because of the optimized gel formulation, a higher signal is achieved for cfDNA smear resulting in a high signal-to-noise ratio and improved assay sensitivity (LOD<25 pg/μL, S/N>3).

A) Electropherogram of the cfDNA sample measured by reference method (competitor method A). B) Stacked Electropherogram obtained for cfDNA sample plated in 3 wells sipped in triplicate in a plate. C) Bar graph representing the average (n=9) concentration of mono-nucleosomal cfDNA measured by LabChip cfDNA assay on different chips and instruments (2 plate cycles) and competitor method A. The average of all LabChip measurements (n=54) is compared with the average of competitor method A (n=9)

In conclusion, we have developed a new assay for the analysis of cfDNA samples. This robust, high-throughput assay can analyze 96 samples in one chip preparation in 1.5 hours. LabChip cfDNA assay offers sensitivity with a higher cfDNA signal smear relative to the competitor method (<25 pg/μL, S/N>3) with a linear dynamic range of 50 pg/μL- 1000 pg/μL for quantitation of cfDNA. Additionally, LabChip cfDNA assay can be used for the analysis of cfDNA samples irrespective of the extraction kits used.

Find the whole application note here.