Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Hohensee, T.
Isolation of circulating tumor cells from patients with metastatic colorectal cancer.
[ Diplomarbeit/Master Thesis ] Karl-Franzens University of Graz; 2013. pp.97.
- Autor*innen der Med Uni Graz:
- Hohensee Tabea
- Betreuer*innen:
- Haybäck Johannes
- Kroneis Thomas
- Altmetrics:
- Abstract:
- CRC is one of the most common cancers worldwide. One fourth of them are diagnosed in an advanced stage, with a shortened survival rate of the effected patients, even after removal of the primary tumor. Based on these facts a better understanding of the formation of metastasis is needed. One approach is the isolation and characterization of circulating tumor cells (CTCs). These cells leave the primary tumor, circulate through the blood- or lymph system and can then settle in another part of the body, forming metastasis. With respect to the OncoTrack project peripheral blood samples of patient suffering from CRC were collected and further processed. This project focuses on finding a more accurate in-silico model for colorectal cancer treatment. There are several techniques available to isolate CTCs focusing on different characteristics, like size, surface antigens or buoyant density of a cell. In this master thesis a method based on negative enrichment with immunomagnetic nanobeads was conducted and improved. While preforming the already established protocol, the remaining leukocytes were not detectable after the antibody staining. So, different antibody combinations were looked at until a better setting was established, followed by whole transcriptome amplification of the isolated single cells or cell pools. Afterwards, these cells were analyzed on a molecular level with TaqMan© probes, using multiplex real-time PCR, which I established in this working group. There, several different tumor markers were tested, containing EpCAM, CK19, CK20, EGFR, PBGD and CEA, and also eIf3a and eIf4e bp2. CD45 was used as a control for leukocyte contamination. During establishment of this method two CRC lines, SW620 and HCT-116 cells were used. Out of 65 amplified patient samples, two could be proven to express keratin 19, indicating that theses isolated cells are likely of CRC origin.