Cow blood - A superior storage option in forensics?

Given the use of modified blood products (e.g. leucocyte depleted erythrocyte concentrates in SAG-mannitol, dehydrated blood powder, defibrinated blood), drawing blood from conscious animals while minimizing their stress is a good option to obtain blood for bloodstain pattern analysis. Nevertheless, the blood must be well described since individual differences in quality can occur, and storage will influence blood components qualitatively and quantitatively. Cow has been discussed as a suitable source of blood supply, but current data lack hematological and full rheological perspectives. This project includes the respective parameters in combination with passive drip pattern experiments during refrigerated storage in multiple study arms. Cow blood displayed a constant increase in viscosity (at high shear rate: 1000s-1), reflecting the expected reduction in red blood cell (RBC) flexibility. RBCs shrank but remained intact with very few irregular shapes, therefore there was no evidence of hemolysis. Influence of storage on stain size in passive drip pattern experiments with different substrates was minimal. However in cows, it is not hemolysis but an early change in suspension properties that indicates storage lesion. Viscosity (at low shear rate: 1s-1) of some blood samples increased three-fold (peaking at day 14), transitioning sharply to near-Newtonian (almost shear-independent) behavior thereafter. The higher this increase in viscosity, the greater the increase in the number of satellite spatter on glass. In order to ensure high quality simulations in the future, comprehensive rheological analyses to detect gradual changes in blood pseudoplasticity should be implemented in the forensic discipline of bloodstain pattern analysis.

Purified Clinoptilolite-Tuff as an Efficient Sorbent for Gluten Derived from Food.

Various gluten-related diseases (celiac disease, wheat allergy, gluten sensitivity) are known and their incidence is growing. Gluten is a specific type of plant storage protein that can impair the health of gluten-prone persons following consumption, depending on the origin. The most severe effects are induced by wheat, barley, and rye. The only treatment is based on the absolute avoidance of those foods, as even traces might have severe effects on human well-being. With the goal of binding gluten impurities after ingestion, an in vitro setting was created. A special processed kind of zeolite, purified clinoptilolite-tuff (PCT), was implemented as an adsorber of gluten derived from different origins. Zeolites are known for their excellent sorption capacities and their applications in humans and animals have been studied for a long time. Tests were also performed in artificial gastric and intestinal fluids, and the adsorption capacity was determined via a certified validated method (ELISA). Depending on the kind of gluten source, 80-130 µg/mg of gluten were bound onto PCT. Hence, purified clinoptilolite-tuff, which was successfully tested for wheat, barley, and rye, proved to be suitable for the adsorption of gluten originating from different kinds of crops. This result might form the basis for an expedient human study in the future.

Animal blood in translational research: How to adjust animal blood viscosity to the human standard.

Animal blood is used in mock circulations or in forensic bloodstain pattern analysis. Blood viscosity is important in these settings as it determines the driving pressure through biomedical devices and the shape of the bloodstain. However, animal blood can never exactly mimic human blood due to erythrocyte properties differing among species. This results in the species-specific shear thinning behavior of blood suspensions, and it is therefore not enough to adjust the hematocrit of an animal blood sample to mimic the behavior of human blood over the entire range of shear rates that are present in the body. In order to optimize experiments that require animal blood, we need models to adapt the blood samples. We here offer mathematical models derived for each species using a multi linear regression approach to describe the influence of shear rate, hematocrit, and temperature on blood viscosity. Results show that pig blood cannot be recommended for experiments at low flow conditions (<200 s-1 ) even though erythrocyte properties are similar in pigs and humans. However, pig blood mimics human blood excellently at high flow condition. Horse blood is unsuitable as experimental model in this regard. For several studied conditions, sheep blood was the closest match to human blood viscosity among the tested species.

Binding and neutralization of C. difficile toxins A and B by purified clinoptilolite-tuff.

Clostridioides difficile (C. difficile) infection is a major public health problem worldwide. The current treatment of C. difficile-associated diarrhea relies on the use of antibacterial agents. However, recurrences are frequent. The main virulence factors of C. difficile are two secreted cytotoxic proteins toxin A and toxin B. Alternative research exploring toxin binding by resins found a reduced rate of recurrence by administration of tolevamer. Hence, binding of exotoxins may be useful in preventing a relapse provided that the adsorbent is innocuous. Here, we examined the toxin binding capacity of G-PUR®, a purified version of natural clinoptilolite-tuff. Our observations showed that the purified clinoptilolite-tuff adsorbed clinically relevant amounts of C. difficile toxins A and B in vitro and neutralized their action in a Caco-2 intestinal model. This conclusion is based on four independent sets of findings: G-PUR® abrogated toxin-induced (i) RAC1 glucosylation, (ii) redistribution of occludin, (iii) rarefaction of the brush border as visualized by scanning electron microscopy and (iv) breakdown of the epithelial barrier recorded by transepithelial electrical resistance monitoring. Finally, we confirmed that the epithelial monolayer tolerated G-PUR® over a wide range of particle densities. Our findings justify the further exploration of purified clinoptilolite-tuff as a safe agent in the treatment and/or prevention of C. difficile-associated diarrhea.

Storability of porcine blood in forensics: How far should we go?

Previous studies on the storability of porcine blood for bloodstain pattern analysis (BPA) focused on abattoir blood only and did not include measurements of viscoelasticity. Although known to provoke echinocyte formation, EDTA is widely used for BPA issues. We compared ageing samples taken from live pigs with abattoir blood and detected considerable differences in hematocrit (HCT), total protein and shear viscosity that even worsened with time. Upon storage, high shear viscosity continuously increased, resulting in a partial loss of the typical shear thinning property of blood. Furthermore, we explored CPDA-1, the gold standard in preserving red blood cells (RBCs), for storage of forensic samples. We found it to be a superior choice for anticoagulation, as the rise of high shear viscosity was attenuated compared to EDTA. When performing oscillation measurements, we found a sudden change of viscoelasticity of blood after 22 days, providing a cut-off for storage time. To highlight the importance of hematological and hemorheological changes upon cold storage, we performed simple drip pattern experiments. These tests revealed a tendency to smaller stain diameters and higher numbers of satellite spatter. While this contradicts expectations from elevated viscosity values, we associate this trend to microscopic inhomogeneities due to storage. We recommend CPDA-1 for prolonged storage of BPA samples and suggest the use of comprehensive test protocols including viscoelasticity for determination of the maximum shelf life of pig blood.