Evaluation of Fluorescein as a Label in Electrochemical and Optical Measurements

Fluorescein is a fluorescent dye, that is commonly used as a fluorescent label in biochemistry, environment protection and ophthalmology. Its fluorescent features are well known, while the knowledge about its electrochemical behavior is still negligible. In our research, we decided to investigate electrochemical activity of fluorescein and determine its concentration with the use of differential pulse voltammetry and spectrofluorimetry in order to compare the utility of each method for fluorescein detection, towards simultaneous optical and electrochemical detection of bioanalytes. Due to the strong effect of pH of the environment on the analyte properties, we utilized buffer solutions of various pH during all experiments. Results of the voltammetric measurements indicated that fluorescein is electrochemically active compound, with the electrochemical activity affected by the pH of environment. Fluorescein undergoes irreversible one-step electrochemical oxidation in the potential range from 0 V to 1.4 V on graphene screen-printed electrodes. Differential pulse voltammetric method allowed to successfully detect fluorescein at micromolar concentration levels. Comparison of the data obtained by each measurement method allowed claiming, that electrochemical methods are more suitable for fluorescein determination in the solutions of low pH, whereas spectrofluorimetric method is more advantageous in the case of alkaline solutions. The further research should be performed in the context of the study on electrochemical activity of fluorescein conjugated with bioanalytes e.g. antibody.