Abstract 5493: A new strategy to assess cytotoxicity using Raman spectroscopy of single isolated nuclei

Abstract

Abstract Single-cell biodetection systems have potential applications in biomedicine and pharmacotherapy, and Raman spectroscopy is an ideal tool for single-cell analysis. Raman spectroscopy biosensors can provide an information-rich “fingerprint” of the overall biochemical composition of single cell undergoing time-dependent changes of cellular biochemistry without reacting with many biologically active compounds. To minimize the interfering signals form cytoplasm, nuclei were isolated from cells for analysis by Raman spectrum to compare the cellar responses and biochemical changes that occur in the nuclei of the HL60 promyelocytic leukemia cell line and drug-resistant cells HL60/ADR after treatment with the antineoplastic drug, doxorubicin. With increasing concentrations of doxorubicin up to 80 nM, the intensities of individual peaks representing nucleic acids and proteins displayed a concentration dependent decrease in the nuclei of HL60 but not HL60/ADR. At 80 nM doxorubicin, there was a 55.6±2.2% (p<0.001) of control in the intensities of Raman peaks representing nuclei acid, but no change of Raman peak intensities of the nuclei of HL60/ADR (96.4±3.1% of control). A significant drop in the intensities of Raman peaks representing nucleic acid was evident around 8 to 10 hours after doxorubicin exposure. Upon incubation with 80nM doxorubicin for 24 hours, thymidine incorporation was 74.2±1.9% and 94±3.8% of control in HL60 and HL60/ADR, respectively and Alamar Blue assay was even less sensitive with the value of 102.0±1.8% and 101.7±2.5% of control in HL60 and HL60/ADR, respectively. To verify the sensitivity and reproducibility of single nuclei Raman spectroscopy, treatment of CCRF-CEM, a T lymphoblastoid cell line, and drug-resistant subline CEM/VBL with anti-microtubule agent, vinblastine was also examined. At 0.4 nM, the intensities of Raman spectrum for nucleic acid dropped to 77.2±1.5% (p<0.001) of control while thymidine incorporation was not significantly inhibited (94.8±4.1% of control). In comparison, CEM/VBL was resistant to vinblastine up to 50 nM with negligible changes in Raman peak intensities of CEM/VBL nuclei (96.8±2.7% of control), and thymidine incorporation was 102.5±1.2% of control. These findings demonstrate that this single nuclei Raman spectroscopy has the ability to detect and identify nuclear changes related to cytotoxicity at lower concentrations and in shorter times than conventional cell based assays. Thus, this new cytotoxicity detection strategy using Raman spectroscopy of single, isolated nuclei may be useful for rapid and sensitive detection of cellular changes in response to chemotherapeutic agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5493.