Development of a Point-of-Care Microfluidic RNA Extraction Slide for Gene Expression Diagnosis after Irradiation

Area of Science:

• Biotechnology and Biomedical Engineering
• Molecular Biology and Genomics
• Radiation Biology and Medical Countermeasures

Background:

• Radiological/nuclear emergencies pose significant threats, necessitating rapid diagnostics for conditions like acute radiation syndrome (ARS).
• Conventional RNA extraction from whole blood is time-consuming, requires specialized equipment, and presents challenges for field deployment.
• Point-of-care diagnostics are crucial for timely medical intervention in emergency scenarios.

Purpose of the Study:

• To miniaturize RNA extraction using a microfluidic-based slide (MBS) for point-of-care diagnostics.
• To evaluate the MBS as a preliminary step towards lab-on-a-chip devices for gene expression (GE) analysis.
• To assess the MBS's utility in predicting hematologic acute radiation syndrome (HARS) severity or identifying RNA microbes.

Main Methods:

• Whole blood samples from healthy donors were irradiated to simulate varying degrees of ARS.
• RNA was extracted using a preliminary microfluidic-based slide (MBS) and compared to conventional column-based (CB) RNA extraction.
• RNA quality, quantity, and gene expression of four radiation-induced genes (FDXR, DDB2, POU2AF1, WNT3) were analyzed using qRT-PCR.

Main Results:

• The MBS yielded twice the total RNA from whole blood compared to the CB method (12.0 ± 5.8 µg vs. 6.6 ± 3.2 µg) in half the extraction time.
• All MBS RNA extracts were DNA-free, whereas 30% of CB extracts were contaminated with DNA.
• Despite lower RNA integrity (RINe) values with MBS, gene expression fold-changes were comparable to the CB method, indicating no significant RNA degradation.

Conclusions:

• The preliminary MBS demonstrates significant advantages, including reduced extraction time, elimination of DNA contamination, and lower blood sample requirement.
• The MBS offers comparable gene expression analysis performance to conventional methods, making it suitable for field applications.
• This microfluidic approach is a promising step towards developing portable lab-on-a-chip devices for rapid, point-of-care diagnostics in radiation emergencies.