P146 – Multiplex Lateral Flow Assay for the Simultaneous Detection of Four Kinds of Body Fluids

15:55

Wednesday August 19^th

Station 16

Duration: 12 minutes

04. Forensic biology

Jinding Liu

The rapid visual multiplex detection of various human body fluids is highly significant for expeditiously resolving criminal cases. Since most commercially available rapid assays are designed for single target based on antigen-antibody binding, they are inefficient for mixed bio-stains evidence. Therefore, developing a method for detecting common human body fluids is crucial for forensic society, especially considering the limited evidence left at a crime scene. This study presents a novel lateral flow biosensor system that integrates multiplex polymerase chain reaction (PCR) with gold-nanoparticle-based immunochromatography for the simultaneous visual identification of four human body fluids (venous blood, semen, saliva, and menstrual blood) by targeting four highly expressed body fluid-specific mRNA biomarkers. The assay employs primers tagged with a set of oligonucleotide sequences and 6-FAM fluorescein during multiplex PCR. The amplicons are then captured on the test strip via hybridization between the barcode and complementary sequences pre-immobilized on the membrane, and are visually detected through a gold-nanoparticle/anti-FAM antibody conjugate. The total assay time, including RNA extraction, reverse transcription, PCR, and strip detection, is approximately 3.5-4 hours, with visual readout within 15 minutes. This method showed high specificity for human body fluid detection. All four human body fluids can be clearly detected at 20× cDNA dilution for unknown samples, and the limit of detection (LOD) reaches 40× for semen, menstrual blood and venous blood. The method successfully detected all target body fluids in fresh samples, as well as in two aged specimens stored at room temperature for more than 6 months. Moreover, the system performed as expected in simulated mixed human body fluid samples, suggesting its potential for analyzing both aged and mixed samples. Overall, this system holds great promise for identifying tissue origin prior to individual identification and can serve as a core platform for detecting various human body fluids.

Authors

Jinding Liu (Shanxi Medical University, China)
Hailing Yang (Shanxi Medical University, China)
Wenrui Tian (Shanxi Medical University, China)
Jiangwei Yan (Shanxi Medical University, China)
Gengqian Zhang (Shanxi Medical University, China)

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P146 – Multiplex Lateral Flow Assay for the Simultaneous Detection of Four Kinds of Body Fluids

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