Acute lymphoblastic leukemia (ALL) is the most common type of malignant neoplasms in the pediatric population. B-cell precursor ALLs (BCP-ALLs) are derived from the progenitors of B lymphocytes. Traditionally, risk factors stratifying therapy in ALL patients included age at diagnosis, initial leukocytosis, and the response to chemotherapy. Currently, treatment intensity is modified according to the presence of specific gene alterations in the leukemic genome. Raman imaging is a promising diagnostic tool, which enables the molecular characterization of cells and differentiation of subtypes of leukemia in clinical samples. This study aimed to characterize and distinguish cells isolated from the bone marrow of patients suffering from three subtypes of BCP-ALL, defined by gene rearrangements, i.e., BCR-ABL1 (Philadelphia-positive, t(9;22)), TEL-AML1 (t(12;21)) and TCF3-PBX1 (t(1;19)), using single-cell Raman imaging combined with multivariate statistical analysis. Spectra collected from clinical samples were compared with single-cell spectra of B-cells collected from healthy donors, constituting the control group. We demonstrated that Raman spectra of normal B cells strongly differ from spectra of their malignant counterparts, especially in the intensity of bands, which can be assigned to nucleic acids. We also showed that the identification of leukemia subtypes could be automated with the use of chemometric methods. Results prove the clinical suitability of Raman imaging for the identification of spectroscopic markers characterizing leukemia cells.

Eosinophils (Eos) play an important role in the immune system’s response releasing several inflammatory factors and contributing to allergic rhinitis, asthma, or atopic dermatitis. Since Eos have a relatively short lifetime after isolation from blood, usually eosinophilic cell line (EoL-1) is used to study mechanisms of their activation and to test therapies. In particular, EoL-1 cells are examined in terms of signalling pathways of the inflammatory response manifested by the presence of lipid bodies (LBs). Here we examined the differences in response to inflammation modelled by various factors, between isolated human eosinophils and EoL-1 cells, as manifested in the number and chemical composition of LBs. The analysis was performed using fluorescence, Raman, and coherent anti-Stokes Raman scattering (CARS) microscopy, which recognised the inflammatory process in the cells, but it is manifested slightly differently depending on the method used. We showed that unstimulated EoL-1 cells, compared to isolated eosinophils, contained more LBs, displayed different nucleus morphology and did not have eosinophilic peroxidase (EPO). In EoL-1 cells stimulated with various proinflammatory agents, including butyric acid (BA), liposaccharide (LPS), or cytokines (IL-1β, TNF-α), an increased production of LBs with a various degree of lipid unsaturation was observed in spontaneous Raman spectra. Furthermore, stimulation of EoL-1 cells resulted in alterations of the LBs morphology. In conclusion, a level of lipid unsaturation and eosinophilic peroxidase as well as LBs distribution among cell population mainly accounted for the biochemistry of eosinophils upon inflammation.

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