Abstract 3280
Background
Colorectal cancer (CRC) is one of the leading cancer with the 55% survival rate. The analysis of tumor microenviroment in fresh surgical samples by flow cytometry focuses on the presence of immune cells. However, no direct access to fresh tumor sample is always possible. Moreover, transport of interesting fresh tumor tissue specimens to distant labs is sometimes neccesary. Therefore, we analyzed both fresh and frozen cells isolated from tumor tissue, stored in hibernation buffer for 1 to 3 days and compared the percentage of EpCAM+, CD45+ and CD3+ cells. We analyzed 26 surgical samples of primary CRC and metastatic samples.
Methods
Surgical samples were obtained from patients with CRC and processed for an isolation of a single cell suspension (SCS) prepared using standard protocol with collagenase. After washing and blocking with mouse serum, cells were stained with fluorescently labeled antibodies (EpCAM-PE, CD45-FITC, CD3-APC). The cells were phenotyped using the FACS ARIAII instrument. Samples were obtained from freshly prepared (n = 19) and frozen cells, stored in DMSO supplemented medium (n = 8). 1 sample was phenotyped both from freshly prepared and frozen cells.
Results
From freshly prepared SCS, the average amount of EpCAM+ and CD45+ cells was 78.7% and 14.7%, respectively. In 12 out of 19 cases we also used anti CD3 antibodies. 2% of infiltrated cells were found to be positive for CD3. When using frozen SCS, we detected a higher positivity for EpCAM (92.85%), CD45 (62%) and CD3 (6%) compared to freshly prepared SCS.
Conclusions
We phenotyped human CRC samples for EpCAM, CD45 and some samples for CD3 marker. We found that for the FACS analysis, the stability of tumor tissue samples seems to be acceptable for isolation of SCS from CRC if stored in hibernation buffer, at 4 °C for 1 to 3 days. When isolated SCS from frozen stocks were used, we detected higher percentages of positive cells for EpCAM, CD45, CD3 markers. Therefore, we do not recommend to compare freshly isolated cells with previously frozen cells in FACS experiments.
Clinical trial identification
Editorial acknowledgement
Legal entity responsible for the study
The authors.
Funding
grants APVV (no.16–0066), VEGA (no. 1/0380/18).
Disclosure
All authors have declared no conflicts of interest.
Resources from the same session
5544 - Evaluation of a radiomic signature of CD8 cells in patients treated with immunotherapy-radiotherapy in three clinical trials.
Presenter: Roger Sun
Session: Poster Display session 3
Resources:
Abstract
1117 - Biomarkers predictive of overall survival in advanced cancer patients treated with a peptide-based cancer vaccine.
Presenter: Shigetaka Suekane
Session: Poster Display session 3
Resources:
Abstract
1922 - Expression of PD-L1 in plasma exosomes of NSCLC patients and its associations with PD-L1 expression of corresponding tumor tissues
Presenter: Shaorong Yu
Session: Poster Display session 3
Resources:
Abstract
5495 - Patient’s perspective on digital biomarkers in advanced urologic malignancies
Presenter: Severin Rodler
Session: Poster Display session 3
Resources:
Abstract
3166 - A comprehensive Pan-cancer study of FGFR Aberrations in Chinese cancer patients
Presenter: Yang Gao
Session: Poster Display session 3
Resources:
Abstract
3277 - A Systemic Inflammation Response Index (SIRI) correlates with survival and could be a Predictive Factor for mFOLFIRINOX in Metastatic Pancreatic Cancer (PC)
Presenter: Vilma Pacheco-Barcia
Session: Poster Display session 3
Resources:
Abstract
2680 - Circulating biomarkers and risk of immune-related adverse events (irAEs) in patients (pts) with advanced Non-small cell lung cancer (aNSCLC) and metastatic melanoma (mMel)
Presenter: Alberto Pavan
Session: Poster Display session 3
Resources:
Abstract
4066 - Breast cancer in young women of Kazakh population depending on germline mutations: results of next-generation sequencing
Presenter: Dilyara Kaidarova
Session: Poster Display session 3
Resources:
Abstract
5514 - Discovery of an ImmunoTranscriptomics signature in blood for early colorectal cancer detection
Presenter: Paolo Angelino
Session: Poster Display session 3
Resources:
Abstract
1595 - Serum Netrin-1 as a Biomarker for Colorectal Cancer Detection
Presenter: Jinzhou Zhu
Session: Poster Display session 3
Resources:
Abstract