Particulate emboli retained by the Intersept (R) transfusion filter; a sem study.

The Intersept (R) transfusion filter is the transfusion filter developed by Johnson and Johnson. Filtration is performed by both adsorption and mechanical retention. Not only microaggregates, but damaged and injured cells are likely to be trapped and/or adsorbed. Such a filter is not injurious to the blood components, but thrombotic material accumulation can occur with fresh blood.     

Purification of Chlamydia trachomatis by a simple and rapid filtration method

A simple method for filter purification of Chlamydia trachomatis from cell culture is described. Crude homogenates of chlamydiae-infected cells were passed through a glass prefilter and a 0.6 microns pore diameter polycarbonate filter. The filtrate was then passed through a 0.2 microns pore diameter filter on which the chlamydiae were trapped. This filter was then back-washed to collect the organisms. These procedures removed cell debris and soluble protein, and yielded particles with a narrow size distribution. The mean yield of viable chlamydiae purified by filtration was 64% when the filters were washed at each stage of the process.     

Effect or preliminary filtration on the functional characteristics of bacterioplankton from Lake Khanka]

The dependence of the functional characteristics of bacterioplankton from the loess of Lake Khanka on the pore size of filtering materials was investigated. Soluble organic matter (SOM), bacteria, and bacterial consumers adsorbed on particles suspended in the lake water were found to filter differently depending on the pore size of the filtering material. Filters with pore size 4.5 microns (filters II) retained up to 20% of SOM and 20-30% of bacterial cells. Filters III with pore size 2.87 microns retained almost 50% SOM and about 40% of bacteria. The double layer of gauze no. 72 (referred to as filter I) with pores size 40 microns was unable to completely retain bacterial consumers. In the case of filtrates I and II, the generation time of bacterioplankton decreased with its increasing average daily concentration. In the case of filtrate III, the generation time of bacterioplankton was minimum and did not depend on its concentration. Oxygen consumption rates per one bacterial cell and per unit biomass in filtrates increased with decreasing pore size of the filters through which they had passed. The bacterial biomass and oxygen consumption rate increased exponentially in filtrates III and logarithmically in filtrates I.     

Blood salvage, proper choice for tumor patients?

Blood transfusion is frequently used in tumor patients, However, allogeneic blood transfusion have been claimed to be associated with an increased risk of negative outcomes, including transfusion reactions, fever, virus transmission, and alloimmunization. Other risks of homologous transfusion specific to the tumor patients are the potential deleterious effects on the recurrence of tumor and indefinite overall survival. On the contrary, autologous blood transfusion offers survival advantages to tumor patients and has been shown to minimize the occurrence of many detrimental allogeneic blood associated effects and complications. It also reduces the volume of banked blood needed and improves the prognosis of patients. However, the quality of salvaged blood is still a matter of debate, because it may contain tumor cells which are associated with potential detrimental effects such as metastasis. So, an advanced technology is needed to remove such contaminants to make autologous blood transfusion safer.     

Blood salvage,proper choice for tumor patients?

Blood transfusion is frequently used in tumor patients, However, allogeneic blood transfusion have been claimed to be associated with an increased risk of negative outcomes, including transfusion reactions, fever, virus transmission, and alloimmunization. Other risks of homologous transfusion specific to the tumor patients are the potential deleterious effects on the recurrence of tumor and indefinite overall survival. On the contrary, autologous blood transfusion offers survival advantages to tumor patients and has been shown to minimize the occurrence of many detrimental allogeneic blood associated effects and complications. It also reduces the volume of banked blood needed and improves the prognosis of patients. However, the quality of salvaged blood is still a matter of debate, because it may contain tumor cells which are associated with potential detrimental effects such as metastasis. So, an advanced technology is needed to remove such contaminants to make autologous blood transfusion safer.     

Clarification of yeast cell suspensions by depth filtration

Depth filtration can be very attractive for initial clarification because of low capital costs and ease of operation. However, there is currently no fundamental understanding of the effects of the filter pore size and morphology on the overall capacity and filtrate quality. The objective of this study was to examine the flux, capacity, and filtrate turbidity of a series of depth filters with different pore size ratings and multilayer structures for the filtration of yeast cell suspensions. Data were analyzed using available fouling models to obtain insights into the flux decline mechanisms. Filters with small pore size provide high filtrate quality at low capacity, with the reverse being true for the larger pore sizes. The multilayer structure of commercial depth filters leads to improved performance, although the choice of layer properties is critical. The highest capacity was achieved using a multilayer filter in which the upper layer allows significant yeast cell penetration into the filter matrix but still protects the retentive layer that is needed for a high quality filtrate.     

Suction sampling technique for obtaining standardized cytologic specimens from the oral mucosa

To enhance the value of exfoliative cytology for the study of the oral mucosa, a simple apparatus was developed to permit adequate sampling of a specific site so that samples collected on different occasions could be compared. The device essentially consists of a collecting cup connected to a blood collection evacuation system. The collecting cup is a modified female half of a stainless-steel filter holder supporting a 13-mm-diameter cellulose filter of 0.05-microns pore size. Suction pressure is applied by means of a 10-mL glass tube of premeasured vacuum. After positioning the collecting cup on the selected site on the buccal mucosa, the vacuum (440 mm Hg) is applied for five seconds. The mucosa is drawn in against the filter, producing a monolayered imprint of cells. This sample may be disengaged from the filter by agitation into a solution; this allows quantitative cytologic studies, such as the measurement of cell numbers by an electronic counter or the estimation of the areas of cells and nuclei by computer-aided image analysis of Cytospin preparations. Five separate samplings from each of three test subjects produced a harvest of 3,000 to 7,000 epithelial cells per sample; the cellular areas ranged from 784 to 1,052 sq microns while the nuclear areas ranged from 18.4 to 21.8 sq microns.     

An on-line method for the reduction of fouling of spin-filters for animal cell perfusion cultures

The main limitation in the use of spin-filters during perfusion cultures of animal cells was revealed to be filter fouling. This phenomenon involves cell-sieve interactions as well as cell attachment to, and growth on, the filter surface. The cell attachment effect has been analysed in the present study during long-term perfusion simulations with CHO animal cells. It was demonstrated that at low filter acceleration, below 6.2 m/s2, a high perfusion rate of 25 cm/h induced rapid filter pore clogging within 3 days, whereas increasing the filter acceleration to 25 m/s2 increased filter longevity from 3 to 25 days, for filters with a pore size of 8.5 microm. Increasing the filter pore size to 14.5 microm improved filter longevity by 84% with less viable and dead cell deposits on the filter surface. However, it was demonstrated that filter longevity was not necessarily dependent on the amount of cell deposit on the filter surface. In the second part of this study, ultrasonic technology was used to reduce filter fouling. Filter vibration, induced by a piezo actuator, improved filter longevity by 113% during CHO cells perfusion cultures.     

A modification of the filtration method for studying the content of nonfilterable cells in erythrocyte suspension

The results of filtration assays provide estimates of the deformability of erythrocytes averaged over the entire suspension. These assays do not distinguish whether the entire population or only its small fraction exhibits abnormal rheological properties. We developed a simple method using a filtrometer to determine the percentage of non-filterable (under given conditions) cells in the erythrocyte suspension. Membrane filters made of a polyethylene terphthalate film had the mean pore diameter of 3.1 microns and the length of cylindrical micropores of 7 microns. The buffer flow rate tb depends on the number of free pores in a filter. The plot of the number of pores clogged by non-filterable cells versus the total number of erythrocytes passed through the filter had a linear portion whose slope represents the relative content Z of non-filterable cells in the suspension. We determined Z for various medium osmolarities u. These data were used to derive the distribution of erythrocytes in ucr, the value of u at which an erythrocyte cannot pass through a pore of a given filter because of geometric limitations. The distribution maximum corresponded to 190-200 mOsm/kg for erythrocytes from the normal blood. This means that normal erythrocytes have the median values of their surface area and area-to-volume ratio of 155-151 microns2 and 1.72-1.68 microns-1, respectively. The half-width of the distribution was approximately 30 mOsm/kg. This finding suggests that the normal blood contains a certain fraction of erythrocytes with a decreased area-to-volume ratio. Our results showed that the distribution is altered in various forms of anemia and in ATP-depleted erythrocyte suspensions.     

Mammalian cell retention in a spinfilter perfusion bioreactor

A spinning cylindrical filter is often used to retain mammalian cells in a continuous perfusion bioreactor. This device, known as a spinfilter, has typically been with pore size smaller than the cell particles (single cells or aggregates) in order to achieve cell separation. For single cells in suspension, such an operation cannot be sustained over a long period of time because of clogging of the filter surface. Recently, screens with openings larger than the average cell size have been used to reduce the incidence of clogging. In this article, we have investigated how the screen size affects cell retention. We also showed why it is necessary to optimize the rotational speed of the spinfilter in order to achieve cell retention and reduce screen clogging. Effects of bulk mixing and perfusion rate on screen fouling cell retention, and cell washout were also investigated. (c) 1992 John Wiley & Sons, Inc.     

Particulate emboli retained by a screen and a depth transfusion filter. A SEM study

Microembolization by platelet leukocyte aggregates is a threat to the pulmonary microvasculature when blood is transfused to patients. Those aggregates can be removed by filters (either depth filter or screen filter); their efficiency, as shown by SFP measurements is not questionable, however they are working in different ways as observed by SEM: mechanical retention and adsorption.     

Choice of Pore Size Can Introduce Artefacts when Filtering Picoeukaryotes for Molecular Biodiversity Studies

Published results of studies based on samples size fractionated by sequential filtration (e.g. 0.2–3 μm) indicate that many ciliate, dinoflagellate and rhizarian phylotypes are found among marine picoeukaryotes. This is somewhat surprising as these protists are typically known as being large organisms (often >10 μm) and no picoplanktonic species have so far been identified. Here, the abundances of ciliate and dinoflagellate phylotypes in published molecular studies of picoeukaryotes are shown to correlate negatively with the pore size chosen for the end filter in the sequential filtrations (i.e. the filter used to collect the microbial biomass). This suggests that extracellular DNA adhering to small particles may be the source of ciliate and dinoflagellate phylotypes in picoplanktonic size fractions. This hypothesis was confirmed using real-time qPCR, which revealed significantly less dinoflagellate 18S rDNA in a 0.8–3-μm size fraction compared to 0.2–3 μm. On average, the abundance of putative extracellular phylotypes decreased by 84–89 % when a 0.8-?μm end filter was used rather than a 0.2-μm end filter. A 0.8-μm filter is, however, not sufficient to retain all picoeukaryotic cells. Thus, selection of filter pore size involves a trade-off between avoiding artefacts generated by extracellular DNA and sampling the entire picoeukaryotic community. In contrast to ciliate and dinoflagellate phylotypes, rhizarian phylotypes in the picoplankton size range do not display a pattern consistent with an extracellular origin. This is likely due to the documented existence of picoplanktonic swarmer cells within this group.     

Picoplankton size distributions in marine and fresh waters: problems with filter fractionation studies

Abstract The retention of algal picoplankton by Nuclepore polycarbonate filters of 0.2, 1.0, 2.0 and 3.0 μm pore size was tested in 2 marine and 3 freshwater sites. When 1 μm Nuclepore filters were used, the percentage of the total cyanobacterial cells passing the filter varied between sites and with increasing depth within sites. As much as 99% of the Synechococcus -like cells was retained by a 1 μm filter. This could lead to an underestimation of the picoplanktonic contribution or, more seriously, an apparent distribution pattern that is an artifact of the choice of filter pore size. Filter retention was also dependent on vaccum pressure during filtration. This study emphasizes the need for direct observation of picoplankton numbers in filter fractionation studies.     

Clarification of recombinant proteins from high cell density mammalian cell culture systems using new improved depth filters

Increasingly high cell density, high product titer cell cultures containing mammalian cells are being used for the production of recombinant proteins. These high productivity cultures are placing a larger burden on traditional downstream clarification and purification operations due to higher product and impurity levels. Controlled flocculation and precipitation of mammalian cell culture suspensions by acidification or using polymeric flocculants have been employed to enhance clarification throughput and downstream filtration operations. While flocculation is quite effective in agglomerating cell debris and process related impurities such as (host cell) proteins and DNA, the resulting suspension is generally not easily separable solely using conventional depth filtration techniques. As a result, centrifugation is often used for clarification of cells and cell debris before filtration, which can limit process configurations and flexibility due to the investment and fixed nature of a centrifuge. To address this challenge, novel depth filter designs were designed which results in improved primary and secondary direct depth filtration of flocculated high cell density mammalian cell cultures systems feeds, thereby providing single‐use clarification solution. A framework is presented here for optimizing the particle size distribution of the mammalian cell culture systems with the pore size distribution of the gradient depth filter using various pre‐treatment conditions resulting in increased depth filter media utilization and improved clarification capacity. Feed conditions were optimized either by acidification or by polymer flocculation which resulted in the increased average feed particle‐size and improvements in throughput with improved depth filters for several mammalian systems. Biotechnol. Bioeng. 2013; 110: 1964–1972. © 2013 Wiley Periodicals, Inc.     

Membrane Filtration of the Reiter Treponeme

A membrane filtration technique with commercially available membrane filters (Millipore Corp.) was effective for the removal of Reiter treponemes from liquids such as fluorescent-antibody conjugates, to which the organisms are added for adsorption. Reiter treponemes from an 8-day culture were not microscopically detectable in filtrates through membranes with a pore diameter of 0.45 μm, but treponemes were demonstrated in the filtrate by cultural methods. No organisms of the 8-day culture passed through a membrane filter having a pore size of 0.22 μm, as determined by microscopy and culture. Culture data indicated that a filter with a pore size of 0.1 μm was necessary to prevent passage of treponemes from 4-day cultures. It is recommended that a membrane filter with a pore size of 0.22 μm or smaller be used for the removal of Reiter treponemes from suspensions and that the age of the culture be considered in choosing filter pore size.     

Penetration of macromolecules into contracted blood clot.

The effectiveness of thrombolytic therapy is determined by accessibility of thrombus compartments to plasminogen activators and, therefore, depends on permeability of thrombus to blood born macromolecules. Accumulation of 125I labeled proteins with molecular massess ranging from 150 to 450 kD into partly contracted blood clot or plasma clot was consistent with diffusion coefficients 3.2 x 10(-11) and 2.7 x 10(-11) m2 s-1, respectively. So far as the model conditions imitated those for venous thrombi, these data indicate that such thrombi are porous enough for immunoconjugates of relatively big size.     

Toward an understanding of the epithelial requirement for osteogenesis in scleral mesenchyme of the embryonic chick

Explants of scleral tissue from chick embryos of H.H. stage 29-36 (6-10 days of incubation) were used to determine if the epithelial-mesenchymal interaction which initiates scleral bone formation is cell contact, extracellular matrix, or diffusion mediated. Transfilter tissue recombinations, in which explanted interacting tissues are associated across interposing Nuclepore filters of various pore sizes and thicknesses, were performed with scleral mesenchyme and epithelium. When filters with pore sizes which would allow the passage of cell processes and diffusible substances were used, osteogenesis was initiated in the scleral mesenchyme. When cell processes were blocked with thicker filters or smaller pore sizes, bone formation still occurred, indicating that a diffusible substance mediates this tissue interaction. Further support for a diffusion-mediated interaction came from transfilter experiments using dialysis membranes to discriminate the size of the molecule(s), and Millipore filters to determine the distance over which these molecules travel. These experiments revealed that the scleral epithelial diffusible factor has a molecular weight of between 3500 and 6000 daltons, and acts over distances between 150 and 300 microns.     

High-density continuous cultures of hybridoma cells in a depth filter perfusion system

A depth filter perfusion system (DFPS) for animal cell culture was developed and its use in continuous highdensity cultures of hybridoma cells was investigated. In the DFPS, based on cell immobilization in a cylindrical depth filter matrix, cells were easily immobilized and cultivated by simple medium recirculation. The cell density in the 20-mum pore size filter matrix reached up to 3 x 10(7) cells/mLin less than 10 days. This resulted in a high monoclonal antibody productivity of 744 mg/L/day, which was 25-35 times higher than that of continuous-suspension cultures using the same cell line. The 20-mum pore filter retained more cells than the 30-mum filterin a shorter period. The DFPS provides advantages of low-cost set-up, easy operation, and scale-up in the cultures of anchorage-independent cells. It also has a high potential for anchorage-dependent cell cultures because of its unusually high surface-to-volume ratio of 450-600 cm(2)/cm(3). (c) 1994 John Wiley & Sons, Inc.     

Occurrence of brain calcifications,iron deposits,and tonic-clonic convulsions following experimental infections with Candida albicans

Summary During the prolonged presence ofCandida albicans in the spleen of Wistar rats, two cases of calcium deposits in the brain were observed. In one case there was one big calcium deposit at some distance from the cortex. In this case, thickening in a limited area of the leptomeninges was found. The animal had generalized tonicclonic convulsions which lead to death. In the second case, calcium deposits formed two separate agglomerations in the posterior part of the thalamus. Serial sections made at six micron intervals showed the presence of calcium deposits to a depth of forty-eight microns. At twenty-four microns other calcium deposits were observed in the thalamus symmetrically located to those described above.Small iron deposits were found in the anterior part of the thalamus and other iron deposits were found in the temporal lobe of the brain. They were arranged in a straight line along the capillary vessels and most of them were seen in the macrophages in the vicinity of the vessels.Since calcium deposits in the brain occurred in the same experimental conditions as reported cases of calcinosis, it is probable that the same mechanism is involved in all these pathological calcium deposits.This work was supported by a Damon Runyon Memorial Fund Grant Nr. 720.     

Influence of the screen material on the fouling of spin filters

Mouse-mouse hybridomas (15 mum mean diameter) were cultivated in a simulated perfusion reactor with spin filter and external recirculation of the medium. Proteins at high concentrations, such as 10% foetal calf serum (FCS), were found to be not responsible by themselves for fouling, even at high recirculation rates. Stainless steel (10 mum pores) in contrast to polyamide (11 mum proes) led to a great accumulation of dead cells and nucleic acids on the screen, finally leading to fouling, as shown by biochemical and microscopic examinations. It is suggested that the high surface charge density of metals compared to polyamide is responsible for attachment of various residues. Stainless steel should rather be replaced by a resistant and nontoxic synthetic material, such as polyamide 66 which was successfully used. FCS should be avoided, since it seems to increased the fouling phenomenon. Moreover, the pore size of the screen should be carefully defined according to the wide size distribution of living and dead cells of the line used (33% of variation of the mean size in our case) as well as fragments. The purpose of the screen being to get rid of fragments and small dead cells, and not to wash too many new small cells, a good retention was achieved here by a 10-mum opening.