Amino‐polyvinyl Alcohol Coated Superparamagnetic Iron Oxide Nanoparticles are Suitable for Monitoring of Human Mesenchymal Stromal Cells In Vivo

Mesenchymal stromal cells (MSCs) are promising candidates in regenerative cell‐therapies. However, optimizing their number and route of delivery remains a critical issue, which can be addressed by monitoring the MSCs’ bio‐distribution in vivo using super‐paramagnetic iron‐oxide nanoparticles (SPIONs). In this study, amino‐polyvinyl alcohol coated (A‐PVA) SPIONs are introduced for cell‐labeling and visualization by magnetic resonance imaging (MRI) of human MSCs. Size and surface charge of A‐PVA‐SPIONs differ depending on their solvent. Under MSC‐labeling conditions, A‐PVA‐SPIONs have a hydrodynamic diameter of 42 ± 2 nm and a negative Zeta potential of 25 ± 5 mV, which enable efficient internalization by MSCs without the need to use transfection agents. Transmission X‐ray microscopy localizes A‐PVA‐SPIONs in intracellular vesicles and as cytosolic single particles. After identifying non‐interfering cell‐assays and determining the delivered and cellular dose, in addition to the administered dose, A‐PVA‐SPIONs are found to be non‐toxic to MSCs and non‐destructive towards their multi‐lineage differentiation potential. Surprisingly, MSC migration is increased. In MRI, A‐PVA‐SPION‐labeled MSCs are successfully visualized in vitro and in vivo. In conclusion, A‐PVA‐SPIONs have no unfavorable influences on MSCs, although it becomes evident how sensitive their functional behavior is towards SPION‐labeling. And A‐PVA‐SPIONs allow MSC‐monitoring in vivo.     

Compact zwitterion-coated iron oxide nanoparticles for biological applications

The potential of superparamagnetic iron oxide nanoparticles (SPIONs) in various biomedical applications, including magnetic resonance imaging (MRI), sensing, and drug delivery, requires that their surface be derivatized to be hydrophilic and biocompatible. We report here the design and synthesis of a compact and water-soluble zwitterionic dopamine sulfonate (ZDS) ligand with strong binding affinity to SPIONs. After ligand exchange, the ZDS-coated SPIONs exhibit small hydrodynamic diameters, and stability with respect to time, pH, and salinity. Furthermore, small ZDS coated SPIONs were found to have a reduced nonspecific affinity (compared to negatively charged SPIONs) toward serum proteins; streptavidin/dye functionalized SPIONs were bioactive and thus specifically targeted biotin receptors.     

LHRH-functionalized superparamagnetic iron oxide nanoparticles for breast cancer targeting and contrast enhancement in MRI

This paper shows that superparamagnetic iron oxide nanoparticles (SPIONs) conjugated to luteinizing hormone releasing hormone (LHRH) (LHRH–SPIONs), can be used to target breast cancer cells. They also act as contrast enhancement agents during the magnetic resonance imaging of breast cancer xenografts. A combination of transmission electron microscopy (TEM) and spectrophotometric analysis was used in our experiments, to investigate the specific accumulation of the functionalized superparamagnetic iron oxide nanoparticles (SPIONs) in cancer cells. The contrast enhancement of conventional T2 images obtained from the tumor tissue and of breast cancer xenograft bearing mice is shown to be much greater than that in saline controls, when the tissues were injected with LHRH–SPIONs. Magnetic anisotropy multi-CRAZED images of tissues extracted from mice injected with SPIONs were also found to have enhanced MRI contrast in breast cancer xenografts and metastases in the lungs.     

Facile synthesis of ultrasmall PEGylated iron oxide nanoparticles for dual-contrast T1- and T2-weighted magnetic resonance imaging

The development of new types of high-performance nanoparticulate MR contrast agents with either positive (T(1)) or dual-contrast (both positive and negative, T(1) + T(2)) ability is of great importance. Here we report a facile synthesis of ultrasmall PEGylated iron oxide nanoparticles for dual-contrast T(1)- and T(2)-weighted MRI. The produced superparamagnetic iron oxide nanoparticles (SPIONs) are of high crystallinity and size uniformity with an average diameter of 5.4 nm, and can be individually dispersed in the physiological buffer with high stability. The SPIONs reveal an impressive saturation magnetization of 94 emu g(-1) Fe(3)O(4), the highest r(1) of 19.7 mM(-1) s(-1) and the lowest r(2)/r(1) ratio of 2.0 at 1.5 T reported so far for PEGylated iron oxide nanoparticles. T(1)- and T(2)-weighted MR images showed that the SPIONs could not only improve surrounding water proton signals in the T(1)-weighted image, but induce significant signal reduction in the T(2)-weighted image. The good contrast effect of the SPIONs as T(1) + T(2) dual-contrast agents might be due to its high magnetization, optimal nanoparticle size for T(1) + T(2) dual-contrast agents, high size monodispersity and excellent colloidal stability. In vitro cell experiments showed that the SPIONs have little effect on HeLa cell viability.     

Superparamagnetic iron oxide nanoparticles assembled magnetic nanobubbles and their application for neural stem cells labeling

Micro/nanobubbles for use as ultrasound contrast agents have been fabricated with different shell materials.When various biomedical nanoparticles have been embedded in the shells of bubbles,the composite structures have shown promising applications in multi-modal imaging,drug/gene delivery,and biomedical sensing.In this study,we developed a new gas-liquid interface self-assembly method to prepare magnetic nanobubbles embedded with superparamagnetic iron oxide nanoparticles(SPIONs).The diameter of the generated assembled nanobubbles was 227.40±87.21 nm with a good polydispersity index(PDI)of 0.29.Under the condition of 150 compression cycles,the nanobubble concentration could reach about 6.12×10⁹/mL.Transmission electron microscopy(TEM)and scanning electronic microscopy(SEM)demonstrated that the assembled nanobubbles had a hollow gas core with SPIONs adsorbed on the surface.Ultrasound(US)imaging and magnetic resonance imaging(MRI)experiments indicated that the assembled magnetic nanobubbles exhibited good US and MR contrast capabilities.Moreover,the assembled magnetic nanobubbles were used to label neural stem cells under ultrasound exposure.After 40 s US exposure,the magnetic nanobubbles could be delivered into cells with 2.80 pg Fe per cell,which could be observed in the intracellular endosome by TEM.Compared with common incubation methods,the ultrasound exposure method did not introduce the potential cytotoxicity of transfection reagents and the efficiency was about twice as high as the efficiency of incubation.Therefore,the assembled magnetic nanobubbles prepared through the pressure-driven gas-liquid interface assembly approach could be a potential US/MRI dual model imaging nanocarrier for regenerative applications.     

Electrosprayed synthesis of red-blood-cell-like particles with dual modality for magnetic resonance and fluorescence imaging

Red blood cells (RBCs) are able to avoid filtration in the spleen to prolong their half-time in the body because of their flexibility and unique shape, or a concave disk with diameter of some 10 μm. In addition, they can flow through capillary blood vessels, which are smaller than the diameter of RBCs, by morphing into a parachute-like shape. In this study, flexible RBC-like polymer particles are synthesized by electrospraying based on electrospinning. Furthermore, magnetite nanoparticles and fluorescent dye are encapsulated in the particles via in situ hydrolysis of an iron-organic compound in the presence of celluloses. The superparamagnetic behavior of the particles is confirmed by low-temperature magnetic measurements. The particles exhibited not only a dark contrast in magnetic resonance imaging (MRI), but also effective fluorescence. The RBC-like particles with flexibility are demonstrated to have a dual-modality for MRI and fluorescence imaging.     

Efficient internalization of peptide-conjugated SPIONs in dendritic cells for tumor targeting

Antigen delivery using nanoparticles becomes useful and novel strategy to develop immunotherapeutic approaches against cancer. In the current study, we examined the feasibility of SPIONs-mediated delivery of antigenic peptides to local tumor for application to cancer immunotherapy. SPIONs carrying murine melanoma antigens, hgp100(25-33) were prepared and used to test its efficacy in mouse model. Efficient uptake of peptide-conjugated SPIONs by murine dendritic cells (DCs) was shown, using NP labeled with the fluorescent dye Furthermore, potential targeting effect of SPIONs carrying tumor antigenic peptide was verified in vitro and in vivo. Our results demonstrate the feasibility of SPIONs-mediated antigen delivery for cancer immunotherapy and highlight the clinical potential of SPIONs for future cancer treatment with high efficacy.     

Characterizing Physical Properties of Superparamagnetic Nanoparticles in Liquid Phase Using Brownian Relaxation

Superparamagnetic iron oxide nanoparticles (SPIONs) have been extensively used as bioimaging contrast agents, heating sources for tumor therapy, and carriers for controlled drug delivery and release to target organs and tissues. These applications require elaborate tuning of the physical and magnetic properties of the SPIONs. The authors present here a search‐coil‐based method to characterize these properties. The nonlinear magnetic response of SPIONs to alternating current magnetic fields induces harmonic signals that contain information of these nanoparticles. By analyzing the phase lag and harmonic ratios in the SPIONs, the authors can predict the saturation magnetization, the average hydrodynamic size, the dominating relaxation processes of SPIONs, and the distinction between single‐ and multicore particles. The numerical simulations reveal that the harmonic ratios are inversely proportional to saturation magnetizations and core diameters of SPIONs, and that the phase lag is dependent on the hydrodynamic volumes of SPIONs, which corroborate the experimental results. Herein, the authors stress the feasibility of using search coils as a method to characterize physical and magnetic properties of SPIONs, which may be applied as building blocks in nanoparticle characterization devices.     

Surface ligand influenced free radical protection of superparamagnetic iron oxide nanoparticles (SPIONs) toward H9c2 cardiac cells

There have been a number of studies which deal with either toxic or non-toxic nature of superparamagnetic iron oxide nanoparticles (SPIONs); however, there is no clear cut information about their exact behavior and the reasons for its dual action. The objective of the present study was to investigate the SPIONs having similar oxidation states, but varying surface ligands and their role in terms of protecting the iron-mediated toxic responses. The four different SPIONs includes: (i) SPIONs containing oleic acid (SPIONs-1), (ii) SPIONs without any surface ligand (SPIONs-2), (iii) SPIONs containing cysteamine ligand (SPIONs-3), and (iv) SPIONs having both of oleic acid and cysteamine ligand. The particle size, surface functionality, and electronic oxidation states were confirmed by the HRTEM, FT-IR, and XPS analysis, respectively. On in vitro testing of all four SPIONs with H9c2 cardiomyocyte cell line, the SPIONs-2 without any surface ligand found to exhibit significant decrease in the viability of cells at a concentration of 200 μg mL^?1 for 16-h exposure period. Further investigation of toxicity mechanism resulted in the fact that the SPIONs-2 involved in the formation of ROS due to the role played by the more electron deficient Fe³⁺ form of iron, there by decreased the glutathione release, increased DNA cleavage, and disrupted the mitochondrial transmembrane potential. However, the presence of unsaturation and/or thiol group (–SH) containing ligands on other SPIONs protected the cardiac cells from undergoing ROS-induced oxidative stress. Further, the results of the study confirming the importance of having unsaturated double bonds and/or –SH group possessing ligands onto the surface of SPIONs by means of protecting the cells from the influence of electron deficient Fe³⁺ state of iron.     

Granzyme B Functionalized Nanoparticles Targeting Membrane Hsp70‐Positive Tumors for Multimodal Cancer Theranostics

Functionalized superparamagnetic iron oxide nanoparticles (SPIONs) have emerged as potential clinical tools for cancer theranostics. Membrane‐bound 70 kDa heat shock protein (mHsp70) is ubiquitously expressed on the cell membrane of various tumor types but not normal cells and therefore provides a tumor‐specific target. The serine protease granzyme B (GrB) that is produced as an effector molecule by activated T and NK cells has been shown to specifically target mHsp70 on tumor cells. Following binding to Hsp70, GrB is rapidly internalized into tumor cells. Herein, it is demonstrated that GrB functionalized SPIONs act as a contrast enhancement agent for magnetic resonance imaging and induce specific tumor cell apoptosis. Combinatorial regimens employing stereotactic radiotherapy and/or magnetic targeting are found to further enhance the therapeutic efficacy of GrB‐SPIONs in different tumor mouse models.     

SPION@APTES@FA-PEG@Usnic Acid Bionanodrug for Cancer Therapy

In this work, we aimed to develop stable usnic acid (UA)-conjugated superparamagnetic iron oxide nanoparticles (SPIONs) as a potential drug carrier for in vitro analysis of MCF-7 (breast cancer cell line), HeLa (cervix cancer cell line), L929 (mouse fibroblast cell line), U87 (glioblastoma cell line, brain cancer), and A549 (human lung cancer cell line) cell lines. SPIONs were synthesized via the polyol method and functionalized with APTES using the Stöber method. Carboxylated polyethylene glycol (PEG-COOH), folic acid (FA), and carboxylated luteolin (CL) were conjugated on the surface via a carboxylic/amine group using the nanoprecipitation method, respectively. X-ray powder diffraction analysis confirmed the purity of the product with crystallite size of around 11 nm. Fourier-transformed infrared spectrophotometer (FT-IR) analyses explained the conjugation of all functional groups to the surface of SPIONs. The percentages of inorganic and organic content in the products were investigated via thermal gravimetric analyzer (TGA). For morphological analysis, a transmission electron microscope (TEM) was used. The superparamagnetic property of the product was also confirmed by vibrating sample magnetometer (VSM).     

New biomimetic constructs for improved in vivo circulation of superparamagnetic nanoparticles

Superparamagnetic iron oxide nanoparticles (SPIOs) have been produced and used as a potent and versatile contrast media for magnetic resonance imaging (MRI). Despite a number of efforts to improve their surface chemistry and biocompatibility, the SPIOs half life in blood circulation is very short and they are rapidly taken up by the reticuloendothelial system (RES). In this paper we describe a new method that permits to avoid the rapid clearance of SPIOs. Nanoparticles are made biocompatible by encapsulation into autologous red blood cells. These biomimetic constructs preserve the main properties of the cells that escape RES clearance as well as the properties of the nanoparticles that perform even better than in blood suspension with reduced T2*. These SPIO-loaded RBCs are promising intravascular imaging contrast agents and could also be addressed to selected body compartments by an external magnetic field.     

In vivo study of anti‐epidermal growth factor receptor antibody‐based iron oxide nanoparticles (anti‐EGFR‐SPIONs) as a novel MR imaging contrast agent for lung cancer (LLC1) cells detection

Superparamagnetic iron oxide nanoparticles (SPIONs) conjugated with anti‐epidermal growth factor receptor monoclonal antibody (anti‐EGFR‐SPIONs) were characterised, and its cytotoxicity effects, ex vivo and in vivo studies on Lewis lung carcinoma (LLC1) cells in C57BL/6 mice were investigated. The broadband at 679.96 cm⁻¹ relates to Fe–O, which verified the formation of the anti‐EGFR‐Mab with SPIONs was obtained by the FTIR. The TEM images showed spherical shape 20 and 80 nm‐sized for nanoparticles and the anti‐EGFR‐SPIONs, respectively. Results of cell viability at 24 h after incubation with different concentrations of nanoprobe showed it has only a 20% reduction in cell viabilities. The synthesised nanoprobe administered by systemic injection into C57BL/6 mice showed good Fe tumour uptake and satisfied image signal intensity under ex vivo and in vivo conditions. A higher concentration of nanoprobe was achieved compared to non‐specific and control, indicating selective delivery of nanoprobe to the tumour. It is concluded that the anti‐EGFR‐SPIONs was found to be as an MR imaging contrast nanoagent for lung cancer (LLC1) cells detection.Inspec keywords: toxicology, biomedical MRI, lung, magnetic particles, biomedical materials, nanofabrication, nanomagnetics, transmission electron microscopy, nanomedicine, superparamagnetism, nanoparticles, iron compounds, proteins, cellular biophysics, molecular biophysics, cancer, tumours, Fourier transform infrared spectraOther keywords: MR imaging contrast agent, LLC1, superparamagnetic iron oxide nanoparticles, Lewis lung carcinoma cells, ex vivo conditions, cell viability, antiepidermal growth factor receptor antibody‐based iron oxide nanoparticles, antiEGFR‐SPION, lung cancer cell detection, antiepidermal growth factor receptor monoclonal antibody, cytotoxicity effects, C57BL‐6 mice, antiEGFR‐Mab, FTIR spectra, TEM, spherical shape, incubation, nanoprobe concentrations, systemic injection, Fe tumour uptake, image signal intensity, in vivo conditions, time 24.0 hour, Fe₃ O₄      

Folate-conjugated zein/Fe<Subscript>3</Subscript>O<Subscript>4</Subscript> nanocomplexes for the enhancement of cellular uptake and cytotoxicity of gefitinib

Integrating various functional components into a single nano-platform is an ideal but challenging strategy for cancer therapy. Herein, a facile approach to fabricating multi-targeted nano-drug delivery systems which can be monitored is reported. Superparamagnetic iron oxide nanoparticles (SPIONs) and gefitinib (GEF) were encapsulated into folic acid-conjugated zein (Fa–zein) nanocomplexes (GEF-FSZs) with good dispersity, high GEF loading efficiency and pH-dependent release profile. The uptake of water-insoluble GEF was facilitated by encapsulating GEF into a zein-based nanocomplex, the magnetic responsive property endowed by SPIONs and the conjugation of Fa, which resulted in enhanced toxicity to A549 cells. The endocytosis study indicated that macropinocytosis and clathrin/caveolae-independent endocytosis exerted great influence on the internalization of GEF-FSZs. These results implied that GEF-FSZs could be a promising candidate for controlled and targeted drug delivery.     

Antibacterial activity of SPIONs versus ferrous and ferric ions under aerobic and anaerobic conditions: a preliminary mechanism study

In modern medicine, major attention has been paid to superparamagnetic iron oxide nanoparticles (SPIONs). Recent studies have shown the antibacterial properties of SPIONs against some Gram‐positive and Gram‐negative bacterial strains. These nanoparticles (NPs) can bind to bacterial membranes via hydrophobic or electrostatic interactions and pass through cell barriers. In this study, the authors evaluated the antibacterial activity of magnetic NPs in comparison with ferrous and ferric ions. The level of reactive oxygen species (ROS) in the treated Staphylococcus aureus and Escherichia coli bacteria were directly measured by fluorometric detection. The results showed that iron ions and SPIONs had significant dependent antimicrobial activities. SPIONs showed greater inhibitory effects than ferrous and ferric ions against the growth of treated bacterial strains under anaerobic conditions, while in aerobic conditions, ferrous showed the strongest antibacterial activity. In anaerobic conditions, they observed the greatest ROS formation and lowest minimum inhibitory concentration in the SPION‐treated group in comparison with the other groups. It seems that the release of iron ions from SPIONs and subsequent activation of ROS pathway are the main antibacterial mechanisms of action. Nevertheless, the greater antibacterial effect of SPIONs in anaerobic conditions represents other mechanisms involved in the antibacterial activity of these NPsInspec keywords: nanomagnetics, antibacterial activity, hydrophobicity, nanoparticles, superparamagnetism, biomedical materials, iron compounds, membranes, nanobiotechnologyOther keywords: ferrous ions, anaerobic conditions, superparamagnetic iron oxide nanoparticles, antibacterial properties, bacterial membranes, electrostatic interactions, bacterial strains, aerobic conditions, SPION‐treated group, antibacterial effect, cell barriers, 2′,7′‐dichlorodihydrofluorescein diacetate, reactive oxygen species, fluorometric detection, Staphylococcus aureus, Escherichia coli     

Superparamagnetic iron oxide nanoparticles (SPIONs): from synthesis to in vivo studies--a summary of the synthesis, characterization, in vitro, and in vivo investigations of SPIONs with particular focus on surface and colloidal properties

In this work, we present a short summary of the synthesis and characterization of superparamagnetic iron oxide nanoparticles and their behavior in vitro and in vivo. Therefore, we have used various characterization techniques to deduce the physical particle size as well as magnetic properties. It is shown that the particle properties were significantly improved by a thermochemical treatment and dialysis, obtaining weakly interacting particles with a clear blocking temperature. We also present the interaction of polyvinyl alcohol and vinyl alcohol/vinyl amine copolymer-coated SPIONs with HELA cells. It is shown that the uptake increased significantly in the presence of a magnetic field and that surface functional groups had an impact on particle uptake and metabolic activity. Furthermore, the influences of the varied parameters (polymer type and therefore surface charge, cell medium, and serum) on the agglomeration rate and the cell uptake are presented and discussed. Finally, we briefly describe the intraarticular application of SPIONs in sheep, their uptake by synovial membrane, and their systemic distribution and elimination.     

Magnetically responsive,sorafenib loaded alginate microspheres for hepatocellular carcinoma treatment

This study aimed to develop sorafenib loaded magnetic microspheres for the treatment of hepatocellular carcinoma. To achieve this goal, superparamagnetic iron oxide nanoparticles (SPIONs) were synthesised and encapsulated in alginate microspheres together with an antineoplastic agent, sorafenib. In the study, firstly SPIONs were synthesised and characterised by dynamic light scattering, energy‐dispersive X‐ray spectroscopy, and scanning electron microscopy. Then, alginate‐SPIONs microspheres were developed, and further characterised by electron spin resonance spectrometer and vibrating sample magnetometer. Besides the magnetic properties of SPIONs, alginate microspheres with SPIONs were also found to have magnetic properties. The potential use of microspheres in hyperthermia treatment was then investigated and an increase of about 4°C in the environment was found out. Drug release studies and cytotoxicity tests were performed after sorafenib was encapsulated into the magnetic microspheres. According to release studies, sorafenib has been released from microspheres for 8 h. Cytotoxicity tests showed that alginate‐SPION‐sorafenib microspheres were highly effective against cancerous cells and promising for cancer therapy.Inspec keywords: drug delivery systems, drugs, nanofabrication, magnetic particles, iron compounds, scanning electron microscopy, hyperthermia, biomedical materials, encapsulation, nanoparticles, light scattering, nanomagnetics, cellular biophysics, toxicology, cancer, nanomedicine, superparamagnetism, nanocomposites, magnetometry, paramagnetic resonance, X‐ray chemical analysisOther keywords: sorafenib loaded alginate microspheres, hepatocellular carcinoma treatment, sorafenib loaded magnetic microspheres, superparamagnetic iron oxide nanoparticles, dynamic light scattering, energy‐dispersive X‐ray spectroscopy, scanning electron microscopy, electron spin resonance spectrometer, vibrating sample magnetometer, hyperthermia treatment, drug release, alginate‐SPION‐sorafenib microspheres, antineoplastic agent, cytotoxicity tests, cancerous cells, time 8.0 hour, Fe₃ O₄      

Preservation of 5300 year old red blood cells in the Iceman

Changes in elasticity and structures of red blood cells (RBCs) are important indicators of disease, and this makes them interesting for medical studies. In forensics, blood analyses represent a crucial part of crime scene investigations. For these reasons, the recovery and analysis of blood cells from ancient tissues is of major interest. In this study, we show that RBCs were preserved in Iceman tissue samples for more than 5000 years. The morphological and molecular composition of the blood corpuscle is verified by atomic force microscope and Raman spectroscopy measurements. The cell size and shape approximated those of healthy, dried, recent RBCs. Raman spectra of the ancient corpuscle revealed bands that are characteristic of haemoglobin. Additional vibrational modes typical for other proteinaceous fragments, possibly fibrin, suggested the formation of a blood clot. The band intensities, however, were approximately an order of magnitude weaker than those of recent RBCs. This fact points to a decrease in the RBC-specific metalloprotein haemoglobin and, thus, to a degradation of the cells. Together, the results show the preservation of RBCs in the 5000 year old mummy tissue and give the first insights into their degradation.     

The Lyophilization of Dispersed Systems: Influence of Freezing Process,Freezing time,Freezing Temperature and RBCs Concentration on RBCs Hemolysis

ABSTRACT

In this work, we studied the influence of different parameters controlling cooling stage on biological dispersed system injury. The human red blood cell (RBCs) was chosen as work model. The study examined the influence of two freezing processes on RBCs hemolysis, one process producing big crystals, the other producing small crystals. Using both processes, we examined the effect of freezing temperature, freezing time, and RBCs concentration on injuries to RBCs. Freezing damage was assessed by the hematocrite measure before freezing and after thawing. The process producing a small number of big ice crystals (Pa) seems—in relation to the one producing a large number of small ice crystals (Pb)—to be less traumatic for the RBC, although the two are not statistically different. Freezing temperature and freezing time influence the preservation of RBCs. At 0 and ?20°C there were high preservation and total hemolysis, respectively. At ?5°C and ?10°C, the RBC hemolysis depends on freezing temperature and freezing time. The RBCs hemolysis rates increases when freezing time increases and when freezing temperature decreases. The rates of RBCs preserved decreases with RBCs concentration some with either the freezing process used (Pa or Pb). More, an accentuation of the difference between the two used freezing processes on RBCs hemolysis was retrieved. The analysis of the conductivity evolution within the RBCs suspension frozen showed that the destruction of the RBCs is had essentially to the solution effects. When the crystallization eutectic takes place, the RBCs are already completely destroyed.     

The lyophilization of dispersed systems: influence of freezing process, freezing time, freezing temperature and RBCs concentration on RBCs hemolysis

In this work, we studied the influence of different parameters controlling cooling stage on biological dispersed system injury. The human red blood cell (RBCs) was chosen as work model. The study examined the influence of two freezing processes on RBCs hemolysis, one process producing big crystals, the other producing small crystals. Using both processes, we examined the effect of freezing temperature, freezing time, and RBCs concentration on injuries to RBCs. Freezing damage was assessed by the hematocrite measure before freezing and after thawing. The process producing a small number of big ice crystals (Pa) seems—in relation to the one producing a large number of small ice crystals (Pb)—to be less traumatic for the RBC, although the two are not statistically different. Freezing temperature and freezing time influence the preservation of RBCs. At 0 and -20°C there were high preservation and total hemolysis, respectively. At -5°C and -10°C, the RBC hemolysis depends on freezing temperature and freezing time. The RBCs hemolysis rates increases when freezing time increases and when freezing temperature decreases. The rates of RBCs preserved decreases with RBCs concentration some with either the freezing process used (Pa or Pb). More, an accentuation of the difference between the two used freezing processes on RBCs hemolysis was retrieved. The analysis of the conductivity evolution within the RBCs suspension frozen showed that the destruction of the RBCs is had essentially to the solution effects. When the crystallization eutectic takes place, the RBCs are already completely destroyed.