Dye-aided optical recognition of reticulocytes and detailed exploration with atomic force microscopy

Reticulocytes are immature red blood cells found in peripheral blood that serve as crucial indicators of bone-marrow function in relation to various pathological conditions. Studies on reticulocytes by means of atomic force microscopy (AFM) are hardly found in the literature, probably due to difficulties regarding their reliable identification and non-invasive scanning with AFM, in their intact form. This study explores both the morphological characteristics and the remnants of the ribosomal ribonucleic acid (rRNA) of intact reticulocytes of healthy donors by means of optical microscopy (OM) aided by a specialized dye, namely brilliant cresyl blue (BCB), and AFM, on a comparative basis. The combination of the advantages of these microscopies enabled us to investigate in detail the overall shape morphology and the structure of the membrane of reticulocytes, in relation to the spatial distribution of residual rRNA. To this end, OM ensures the reliable identification of reticulocytes through the residual rRNA stained by BCB, while AFM provides morphological information down to the nanoscopic level. Our combined OM and AFM results constitute an important contribution to the very limited current knowledge existing in the literature on the spatial distribution of rRNA in reticulocytes found in the peripheral blood of healthy donors.