Biology subjects

Dehghani, M.

Publications and source records attributed to Dehghani, M..

2 recordsLinked to original sources

bioRxiv2019

Tangential flow microfluidics for the capture and release of nanoparticles and extracellular vesicles on conventional and ultrathin membranes

Membranes have been used extensively for the purification and separation of biological species. A persistent challenge is the purification of species from concentrated feed solutions such as extracellular vesicles (EVs) from biological fluids. We investigated a new method to isolate micro- and nano-scale species termed tangential flow for analyte capture (TFAC), which is an extension of traditional tangential flow filtration (TFF). Initially, EV purification from plasma on ultrathin nanomembranes was compared between both normal flow filtration (NFF) and TFF. NFF resulted in rapid formation of a protein cake which completely obscured any captured EVs and also prevented further transport across the membrane. On the other hand, TFF showed capture of CD63 positive EVs with minimal contamination. We explored the use of TFF to capture target species over membrane pores, wash and then release in a physical process that does not rely upon affinity or chemical interactions. This process of TFAC was studied with model particles on both ultrathin nanomembranes and conventional thickness membranes (polycarbonate track-etch). Successful capture and release of model particles was observed using both membranes. Ultrathin nanomembranes showed higher efficiency of capture and release with significantly lower pressures indicating that ultrathin nanomembranes are well-suited for TFAC of delicate nanoscale particles such as EVs.

bioengineering↗

bioRxiv2019

Exosome labeling by lipophilic dye PKH26 results in significant increase in vesicle size

Exosomes are membrane vesicles secreted by cells and distributed widely in all biofluids. Exosomes can modulate the biological activities of cells in a paracrine or endocrine manner, in part by transferring their content, such as miRNA, following uptake in recipient cells. Fluorescent labelling of exosomes is a commonly used technique for understanding their cellular targeting and biodistribution. Lipophilic fluorescent dyes such as those in the PKH family have been widely used for exosome labelling. One concern with the use of lipophilic dyes is an increase in the exosome size due to membrane fusion or intercalation. This size shift alone may undermine the validity of exosomes tracing studies as small changes in the size of inorganic nanoparticles are known to affect their cellular uptake and biodistribution. Here, the possibility of minimizing the size shift of PKH labelled exosomes was systematically studied by changing the labelling condition. Unfortunately, the size shift towards larger particles was observed in all the PKH labelling conditions, including those where the labelled exosomes were below the fluorescent detection limit. As opposed to lipophilic dyes, no significant shift in the size of labelled exosomes was detected with protein binding dyes. Since the size shifts identified in all the PKH labelling conditions are likely to affect the cellular uptake and biodistribution, PKH may not be a reliable technique for exosomes tracking.

biochemistry↗