Establishment of an advanced flow cytometry protocol for analyzing immune cell subsets from lung cancer patients

Abstract

Lymphocytes are known for their role in tumor promotion and tumor suppression. T cell infiltration of the tumor microenvironment has been linked to a better prognosis and outcome in various malignancies. CD8+ T cells can eliminate cancer cells by causing them to lyse, while CD4+ T cells can be both tumor promoting and tumor suppressing. However, the prognostic impact of functional lymphocyte subtypes in non-small cell lung cancer (NSCLC) is not completely known. Therefore, the aim of this thesis was to establish a multiparametric flow cytometry protocol to explore the presence of functional lymphocyte subtypes in peripheral blood mononuclear cells from healthy donors and validate it for use in blood and tissue samples from NSCLC patients. Furthermore, the effects of cryopreservation on sample quality and lymphocyte subtype distribution were also explored. Our study showed that the presence of selected lymphocyte subtypes can be explored in blood from NSCLC patients. However, it also showed that cryopreservation had an effect on the distribution of some lymphocyte subtypes. The blood samples that were analyzed using the CD8+ T cell subtypes panel showed a quantitative decrease of cells after cryopreservation. Similar observations were made for blood samples analyzed using the expanded lymphocyte panel. Statistical analysis showed that these differences may be attributable to the effects of cryopreservation. However, due to small sample sizes we were unable to determine whether the differences were statistically significance. Having established this protocol, our future aim is to analyze more NSCLC patient samples, both from blood and tissue.