Non-enriched comparisons can vary, but typically would include grouped animals in standard laboratory housing without objects and with stable food and water locations. Enrichment is a safe non-pharamacological tool to examine behavioral and neurobiological processes in animal models of the lifespan, brain dysfunction and injury.Milk is a complex fluid that contains various types of proteins and extracellular vesicles (EVs). Some proteins can mingle with EVs, and interfere with their isolation. Among these proteins, caseins form micelles of a size comparable to milk EVs, and can thus be co-isolated with EVs. Preliminary steps that affect milk are crucial for EV isolation and impact the purity and abundance of isolated EVs. In the course of our previous works on cow's milk EVs, we found that sodium citrate (1% final), which is a biocompatible reagent capable of breaking down casein micelles into 40-nm monomers, allowed the isolation of high quantities of EVs with low coprecipitation of caseins or other contaminating proteins. Using this protocol, we successfully separated different EV subsets, characterized in depth their morphology, protein content and small RNA enrichment patterns. We were also able to describe their biological function in a mouse model of intestinal inflammation. We, hereby, detail the differential ultracentrifugation procedure that leads to high quantify, medium specificity, isolation of different milk EV subsets from the same sample. More specifically, we highlight the use of sodium citrate as a standardized approach to isolate and study milk EVs and its potential for isolation techniques other than differential ultracentrifugation.Exosomes, a subtype of extracellular vesicles, are nanovesicles of endocytic origin. Exosomes contain a plethora of proteins, lipids, and genetic materials of parent cells to facilitate intercellular communications. Tracking exosomes in vivo is fundamentally important to understand their biodistribution pattern and the mechanism of biological actions in experimental models. Until now, a number of tracking protocols have been developed, including fluorescence labeling, bioluminescence imaging, magnetic resonance imaging, and computed tomography (CT) tracking of exosomes. Recently, we have shown the tracking and quantification of exosomes in a spinal cord injury model, by using two tracking approaches. More specifically, following intranasal administration of gold nanoparticle-encapsulated exosomes to rats bearing complete spinal cord injury, exosomes in the whole central nervous system were tracked by using microCT, and quantified by using inductively coupled plasma and flame atomic absorption spectroscopy. BAL-0028 In addition, optical imaging of fluorescently labeled exosomes was performed to understand the abundance of migrating exosomes in the spinal cord lesion, as compared to the healthy controls, and to further examine their affinity to different cell types in the lesion. Thus, the protocol presented here aids in the study of exosome biodistribution at both cellular and organ levels, in the context of spinal cord injury. This protocol will also enable researchers to better elucidate the fate of administered exosomes in other models of interest.We present a safe and low-cost method suitable for DNA extraction from mycelium and tree tissue samples. After sample preparation, the extraction takes about 60 min. Method performance was tested by extracting DNA from various tree tissue samples and from mycelium grown on solid and liquid media. DNA was extracted from juvenile and mature host material (Picea abies, Populus trichocarpa, Pseudotsuga menziesii) infected with different pathogens (Heterobasidion annosum, Heterobasidion parviporum, Leptographium wagenerii, Sphaerulina musiva). Additionally, DNA was extracted from pure cultures of the pathogens and several endophytic fungi. PCR success rate was 100% for young poplar material and fungal samples, and 48-72% for conifer and mature broadleaved plant samples. We recommend using 10-50 mg of fresh sample for the best results. The method offers a safe and low-cost DNA extraction alternative to study tree-fungus interactions, and is a potential resource for teaching purposes.Parasites of the genus Leishmania infect the mammalian hosts, including mice and humans and cause cutaneous or visceral leishmaniasis depending upon the parasite species transmitted by the vector sandfly. Leishmania amazonensis is one of the Leishmania species responsible for the cutaneous form of the disease. We have inoculated with these parasites the ear dermis of mice. RNA preparations were performed from fragmented tissues using a buffer containing guanidin isothiocynate (RLT buffer, RNeasy Mini Kit, Qiagen, SAS, France) and β-mercaptoethanol. Both reagents facilitate the isolation of intact RNA from tissues and the use of the RNeasy Kits present with several advantages that facilitate the isolation of pure non-degraded total RNA i) This method allows to avoid the presence of phenol in the RNA extraction buffer, commonly used in alternative protocols; ii) Moreover Diethylpyrocarbonate (DEPC) treatment of glassware, to avoid RNAses contamination of the samples, is not required with this protocol; iii) Finally, it is a fast procedure and the isolated total RNA may be concentrated in a small volume thus facilitating its use for downstream experimental procedures.As obesity becomes a global epidemic, the metabolism research field is increasingly focusing on studying the physiological and pathological roles of adipose tissues (AT). However, extracting proteins from AT is challenging due to abundant fat content of intracellular lipid droplets. Several commercial kits for extraction of AT proteins are available, as are protocols (such as the RELi protocol as well as other protein precipitation protocols). The protocols have been introduced to improve the quality and yield of extractions, but these methods either increase the cost or involve multiple steps. Herein, we describe a detailed protocol for mouse AT protein extractions based on our daily laboratory practice. This protocol requires only very common reagents and instruments, and can be completed in 90-120 min and provides good recovery of total protein content. Thus, this protocol is an economically attractive, time-saving and efficient way to extract proteins from the AT.