Alginate/xanthan gum hydrogels as forensic blood substitutes for bloodstain formation and analysis

Understanding the behaviour of human blood outside of the body has important implications in forensic research, especially related to bloodstain pattern analysis (BPA). The design of forensic blood substitutes (FBSs) can provide many advantages, including the incorporation of multiple physiological components for use as safe and reliable materials for forensic applications. In this work, we present the design of synthetic alginate and xanthan gum-based hydrogels that contain electrosprayed microparticles (MPs) with and without crosslinked DNA. In addition to the MPs, the alginate/xanthan gum FBS materials include fillers to alter the physical appearance and fluid properties of the material. The optimized FBS consisted of alginate (1% w/v) and xanthan gum (5.0 × 10^-3% w/v), 2 mM CaCl₂, ferric citrate (0.5% w/v), magnesium silicate (0.25% w/v), Allura Red dye (2% w/v), 0.025% v/v Tween 20 and 9.5% v/v MPs. The FBS was tested in passive dripping experiments relevant to BPA scenarios at various impact angles. The spreading ratio (D_s/D₀) was determined for 90° stains made on a paper surface and compared to bovine blood where the FBS was shown to simulate accurate and predictable spreading behaviour. In addition, we simulated other common BPA scenarios (e.g., impact patterns) and evidence processing potential. The FBS could be swabbed, and the DNA could be extracted, amplified, and genotyped analogous to human blood evidence. A stability test was also conducted which revealed a shelf-life of over 4 weeks where the material remains relevant to human blood at physiological temperature.