Microfluidic Isolation of Aptamers for Intracellular Measurement of Radio-Responsive Proteins.

In large-scale radiological events, there is a need to triage affected individuals based on their biological absorbed dose. Biodosimetry measures biological responses in relation to the received dose. Radiation-responsive protein biomarkers in peripheral blood lymphocytes, especially intracellular proteins, have been validated for biodosimetry with immunochemical-based measurement methods. However, these antibody-based assays can suffer from stability and batch-to-batch variations. Aptamers are single-stranded oligonucleotide alternatives to antibodies that are stable and much smaller in size, making them ideal probes for intracellular targets. However, few aptamers have been developed against intracellular targets, and these efforts are especially hampered due to the time-consuming nature of the conventional aptamer selection method. An efficient method for isolating aptamers against intracellular radiation-responsive proteins is not available yet. Herein, we used a microfluidic aptamer isolation method to develop an aptamer against the intracellular radiation biomarker BAX in blood lymphocytes. The isolated aptamer has a dissociation constant of 6.95 nM against human BAX protein and a bright detail similarity score of 1.9 when colocalizing with anti-BAX aptamer intracellularly. The in situ labeling of the intracellular BAX protein also shows the aptamer can be used to differentiate 2.5 Gy or 3 Gy of radiation in ex vivo human and in vivo mouse peripheral blood samples exposed to X-rays. In conclusion, this proof-of-concept study indicates that the microfluidic-enabled aptamer isolation method could be used for the development of a panel of targeted intracellular proteins for radiation biodosimetry applications.