Labeling Of Pk-15 Cell Line with Nanoparticles of Hadolinium Orthovanadate: Influence of time and Incubation Conditions

Currently, nanoparticles are widely used in medicine and biology. In particular, it was shown that the accumulation of nanoparticles in cells depended on many factors: chemical composition, shape, size as well as composition of the incubation solution and the type of cells interacting with nanoparticles. Based on this, the PK-15 cell line was studied due to constitutively expressing Gal-α-1,3-Gal xenoantigen (α-Gal-epitope) on the cell membrane. This epitope activates the reaction of hyperacute rejection at transplantation; therefore the question of its elimination is crucial when using bioimplants, valve prostheses, bioscaffolds, which originate from animal tissues. Elimination of this epitope or reduction of it expression on the cell surface may prolong graft survival. Thus, the study of the possibility of using nanoparticles to modify the surface of Gal-α-1,3-Gal-bearing cells is relevant. The purpose of the study was to determine the main parameters of PK-15 cell labeling with nanoparticles based on gadolinium europium orthovanadate GdYVO4:Eu3+ and GdVO4:Eu3+ (incubation time, temperature and composition of the solution, need for shaking). Material and methods. Using cell culture techniques and fluorescence microscopy, the labeling of the PK-15 cell line with nanoparticles was studied under various conditions: nanoparticle concentration, incubation time, incubation temperature, introduction of 10% fetal calf serum to the incubation solution, shaking. Different in size and shape nanoparticles based on orthovanadate gadolinium europium were used in the experiment: spherical GdYVO4: Eu3+ (1-2 nm) and spindle-shaped GdVO4: Eu3+ (25×8 nm). Results and discussion. The study results showed that incubation of cells with nanoparticles for 60 minutes at 22°C was sufficient for binding nanoparticles of various shapes and sizes with PK-15 cells. At 0°C incubation conditions, shaking was necessary to facilitate the contact of cells with nanoparticles. The use of incubation solutions containing fetal calf serum turned out to be inexpedient, since under these conditions no nanoparticle fluorescence wass observed. This may be due to the fact that fetal calf serum prevents the interaction of nanoparticles with cell membranes. The prospect of the research is a more detailed study of the interaction of nanoparticles with the membrane of cells enriched with the Gal-α-1,3-Gal epitope. Particularly relevant is the study of the possibility of its deactivation or masking by binding to nanoparticles. Conclusions. At a temperature of 22 ° C for interaction of cells of the line РК-15 from the LF 60 minutes is sufficient time for incubation, while constant shaking is irrelevant. When using the incubation time of 60 minutes at 0 ° C, shaking is necessary to facilitate the contact of cells from the LF. The use of incubation solutions containing 10% fetal calf serum is not feasible, since under these conditions there is no fluorescence of the LF that binds to the cells.