| Autor: |
Stiekema, J., Cats, A., Boot, H., Langers, A. M. J., Balague Ponz, O., Velthuysen, M. L. F., Braaf, L. M., Nieuwland, M., Sandick, J. W. |
| Předmět: |
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| Zdroj: |
Diseases of the Esophagus; Nov/Dec2016, Vol. 29 Issue 8, p1100-1106, 7p |
| Abstrakt: |
The process of preparing endoscopic esophageal adenocarcinoma samples for next-generation DNA/ RNA sequencing is poorly described. Therefore, we assessed the feasibility and pitfalls of preparing esophageal adenocarcinoma endoscopic biopsies toward DNA/ RNA samples suitable for next-generation sequencing. In this prospective study, four tumor biopsy samples were collected from consecutive esophageal cancer patients during esophagogastroduodenoscopy and fresh-frozen in liquid nitrogen. DNA and RNA were isolated from samples with a tumor percentage of at least 50%. For next-generation sequencing, double-stranded DNA (ds DNA) is required and high-quality RNA preferred. The quantity ds DNA and RNA quantity and quality were assessed with the Nanodrop 2000 spectrophotometer ( Thermo Fisher Scientific, Waltham, MA, USA) and Agilent 2100 Bioanalyzer ( Agilent, Santa Clara, CA, USA). Biopsy samples of 69 consecutive patients with esophageal adenocarcinoma were included. In five patients (7%), the tumor percentage was less than 50% in all four biopsies. Using a protocol allowing simultaneous DNA and RNA isolation, the median dsDNA yield was 2.4 μg (range 0.1-12.0 μg) and the median RNA yield was 0.5 μg (range 0.01-2.05 μg). The median RNA integrity number of samples that were fresh-frozen within 30 minutes after sampling was 6.7 (range 4.2-8.9) compared with 2.5 (1.8-4.5) for samples that were fresh-frozen after 2 hours. The results from this study show that obtaining ds DNA and RNA for next-generation sequencing from endoscopic esophageal adenocarcinoma samples is feasible. Tumor percentage and ds DNA/ RNA yield and quality emphasize the need for sampling multiple biopsies and minimizing the delay before fresh-freezing. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
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