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PubMed2004

Sampling Saccharomyces cerevisiae cells by rapid filtration improves the yield of mRNAs.

To optimize the recovery of mRNAs extracted from yeast, different methods for sampling the yeast cells have been compared. For Saccharomyces cerevisiae strains growing on gluconeogenic carbon sources (derepressed cells) rapid filtration allowed much higher yields (3-10 fold) than centrifugation at room temperature or at 4 degrees C. Recovery of total RNA was similar with the different procedures. For S. cerevisiae growing on glucose, filtration caused a 2-4 fold improvement on the mRNA yields obtained from cells sampled by centrifugation. It was also observed that, when derepressed cells of S. cerevisiae W303-1A were collected by filtration and flash-frozen, part of the 25S and 18S rRNAs (up to 50%) was recovered in an unprocessed 32S or 33S form.

Base Sequence↗
PubMed2005

A gel filtration assay to determine glycogen synthase activity.

We developed a gel filtration assay for the determination of glycogen synthase activity in cultured cells or tissue homogenates. Compared to the commonly used filter paper assay, the gel filtration assay resulted in a more than 5-fold reduction of background levels leading to an--at least--twofold increase in precision. These benefits allow the gel filtration method to detect differences of +/-5% in enzyme activity out of 300 microg total cell protein. In addition to high precision and sensitivity, the method's additional salient advantages include lesser expenditure of time and labour and reduced exposure time of the personnel to radioactivity.

Animals↗
PubMed2004

Effect of electrostatic, hydrodynamic, and Brownian forces on particle trajectories and sieving in normal flow filtration.

Particle deposition and fouling are critical factors governing the performance of microfiltration and ultrafiltration systems. Particle trajectories were evaluated by numerical integration of the Langevin equation, accounting for the combined effects of electrostatic repulsion, enhanced hydrodynamic drag, and Brownian diffusion. In the absence of Brownian forces, particles are unable to enter the membrane pores unless the drag associated with the filtration velocity can overcome the electrostatic repulsion. Brownian forces significantly alter this behavior, allowing some particles to enter the pore even at low filtration velocities. The average particle transmission, evaluated from the probability of having a particle enter the pore, increases with increasing filtration velocity due to the greater hydrodynamic drag force on the particle. These results provide important insights into particle behavior in membrane systems.

Journal Article↗
PubMed2005

Experimental aspects of ultrasonically enhanced cross-flow membrane filtration of industrial wastewater.

Ultrasound based on-line cleaning for membrane filtration of industrial wastewater was studied. An ultrasonic transducer was assembled in the membrane module in order to get an efficient cleaning of membranes in fouling conditions. The focus of the studies was on the effects of the ultrasound propagation direction and frequency as well as the transmembrane pressure. The more open the membrane was the easier the membrane became plugged by wastewater colloids, when the ultrasound propagation direction was from the feed flow side of the membrane. If the membrane was tight enough, the ultrasound irradiated from the feed side of the membrane increased the flux significantly. However, in the circumstances studied, the power intensity needed during filtration was so high that the membranes eroded gradually at some spots of the membrane surface. It was discovered that the ultrasonic field produced by the used transducers was uneven in pressurised conditions. On the other hand, the ultrasound treatment at atmospheric pressure during an intermission pause in filtration turned out to be an efficient and, at the same time, a gentle method in membrane cleaning. The input power of 120 W (power intensity of 1.1 W/cm2) for a few seconds was sufficient for cleaning. The flux improvement was significant when using a frequency of 27 kHz but only minor when using 200 kHz.

Equipment Failure↗
PubMed2001

Peritubular filtration as cause of severe hypotony after Ahmed valve implantation for glaucoma.

PURPOSE: To report peritubular filtration as the cause of severe hypotony after Ahmed valve implantation. METHODS: Interventional case report. In a 65-year-old man with primary open-angle glaucoma, ultrasound biomicroscopy was used to image the peritubular region of his right eye that had severe hypotony during the early postoperative period after an Ahmed valve implantation. RESULTS: The ultrasound biomicroscopy examination showed peritubular filtration. Additional surgery consisting of tube extraction, closure of the previous insertion opening, reformation of the anterior chamber with viscoelastic, and reintroduction of the tube through a new paracentesis was performed to solve this complication. Twenty-four hours postoperatively, the intraocular pressure increased to 10 mm Hg. During the 6-month follow-up, the mean intraocular pressure was 16.36 mm Hg (standard deviation, 1.45). CONCLUSION: The possibility of peritubular filtration should be considered in eyes with severe hypotony after restrictive valve implantation.

Aged↗
PubMed2001

Risk factors for failure of 5-fluorouracil needling revision for failed conjunctival filtration blebs.

PURPOSE: To investigate the risk factors for failure of 5-fluorouracil (5-FU) needling revision, a useful procedure for restoring a failed filtration bleb. DESIGN: Interventional case series. METHODS: Retrospectively conducted study. SETTING: Institutional. STUDY POPULATION: Sixty-four eyes of 64 consecutive glaucoma patients that underwent 5-FU needling revisions for failed filtering bleb following either trabeculectomy or phaco-trabeculectomy with or without adjunctive mitomycin C (MMC). OBSERVATION PROCEDURES: Goldmann applanation tonometry, Kaplan-Meier survival analysis, and Cox proportional hazards regression analysis. MAIN OUTCOME MEASURES: Successful outcome of the initial 5-FU needling revision, arbitrarily defined as target intraocular pressure (IOP) control with not more than two topical glaucoma medications and no additional 5-FU needling or other surgical procedures, was analyzed by Kaplan-Meier survival analysis, and risk factors for failure of the initial 5-FU needling revision were analyzed by Cox proportional hazards regression analysis. RESULTS: The cumulative success rate of the initial 5-FU needling revision was 45% at 1 year, 33% at 2 years, and 28% at 4 years. Failure of the initial 5-FU revision correlated significantly with preneedling IOP > 30 mm Hg (P =.0003), lack of MMC use during the previous filtration surgery (P =.013), and IOP >10 mm Hg immediately following needling revision (P =.0012) according to Cox's proportional hazards regression analysis. CONCLUSIONS: Pre-needling IOP > 30 mm Hg, lack of MMC use during the previous filtration surgery, and IOP > 10 mm Hg immediately after needling were found to be significant risk factors for failure of the initial 5-FU needling procedure. Therefore, it is important to monitor IOP closely following needling revision in those patients with such risk factors. They are more likely to require additional therapeutic interventions, including repeat needling revisions.

Aged↗
PubMed1998

Local effects of previous conjunctival incisional surgery and the subsequent outcome of filtration surgery.

PURPOSE: Previous ocular surgery involving conjunctival incision is a risk factor for failure of filtration surgery. To determine whether using adjunctive antimetabolite therapy may have a rational basis, a study was performed to determine whether the conjunctival cellular profile was altered by such surgery. METHODS: After a mean follow-up of 5.9 years, filtration surgery outcomes for 35 patients who had undergone conjunctival incisional surgery were retrospectively compared with 29 control and 18 matched patients who had not undergone such surgery. At the time of filtration surgery, a conjunctival biopsy specimen was obtained from all 82 patients, and these were quantitatively analyzed by light microscopy in a prospective manner. RESULTS: Trabeculectomy success for the control group (93%) was significantly higher than for the previous surgery group (38%) (P < .001). Compared with control tissue, conjunctiva from the patients who had undergone previous surgery contained more fibroblasts (P < .001, P < .05), macrophages (P < .01, P < .001), and lymphocytes (P = .001, P < .01) in both superficial and deep substantia propria (respective P values). Furthermore, trabeculectomy failure was associated with an increase in number of conjunctival fibroblasts in intraoperative specimens. CONCLUSIONS: Previous ocular surgery involving the conjunctiva increases the number of conjunctival fibroblasts and inflammatory cells. This may account for the increased risk of trabeculectomy failure. Perhaps on this basis, the use of adjunctive medication may be justified in patients who have undergone previous ocular surgery involving conjunctival incision.

Adolescent↗
PubMed1983

Gel filtration in sodium dodecyl sulphate of hydrophobic muscle proteins on Sephacryl S-400 Superfine.

Gel filtration has been applied to muscle proteins in the presence of the detergent sodium dodecyl sulphate (SDS). This detergent not only solubilizes a variety of proteins but also increases their hydrodynamic volumes, which is a disadvantage for the process of gel filtration. Nevertheless, appropriate choice of gel type, in this case Sephacryl S-400 Superfine, and optimal elution conditions make SDS gel filtration a useful tool in the separation of heterogeneous protein mixtures in the molecular weight range of 20,000-200,000.

Acrylic Resins↗
PubMed1984

Isolation of detergent-extracted Sendai virus proteins by gel-filtration, ion-exchange and reversed-phase high-performance liquid chromatography and the effect on immunological activity.

Virus envelope proteins were isolated from Triton X-100 extracts of purified Sendai virions by gel-filtration, ion-exchange and reversed-phase high-performance liquid chromatography (HPLC). The fusion protein F, the matrix protein M and the tetrameric and dimeric form of the HN protein were isolated by gel-filtration HPLC with a solvent containing 0.1% sodium dodecyl sulphate. HN and F were also isolated by ion-exchange HPLC with 0.1% Triton X-100 in the eluent. Reversed-phase HPLC was performed on a C1 column with acetonitrile as the organic solvent. Especially the F1 and F2 component of the fusion protein F were obtained in pure form. The immunological activity of the proteins after HPLC was determined with an enzyme-linked immunosorbent assay (ELISA). After gel-filtration and ion-exchange HPLC, proteins still reacted with antiserum to the intact virus while proteins purified by reversed-phase HPLC did not react.

Animals↗
PubMed1998

Separation of motile spermatozoa from frozen-thawed buffalo semen: swim-up vs filtration procedures.

Three experiments were conducted to maximize the recovery rate of motile spermatozoa from frozen-thawed buffalo semen. In Experiment 1, the swim-up of motile spermatozoa was performed in the presence or absence of HEPES in TALP medium and CO2 in the environment. The recovery rate of motile spermatozoa in TALP medium (control), TALP + HEPES + CO2, TALP + HEPES and TALP + CO2 was 15, 18, 12 and 10%, respectively (P > 0.05), with sperm motility at 87, 89, 90 and 90%, respectively (P > 0.05). In Experiment 2, the pH of TALP medium was adjusted to 7.0, 7.5, 8.0, 8.5 and 9.0, and swim-up procedure was performed in the presence of HEPES and CO2. The recovery rate of motile spermatozoa at different pH was 14, 20, 24, 27 and 16%, respectively (P < 0.05). Motility of separated spermatozoa was 88, 91, 90, 89 and 90%, respectively (P > 0.05). In Experiment 3, the efficiency of ion-exchange filtration and Swim-up procedure in separating motile spermatozoa from frozen-thawed buffalo semen was compared. The recovery rate of motile spermatozoa was 95% in filtration procedure and 33% in swim-up procedure (P < 0.005). In all experiments, normal acrosomes did not vary due to treatments (P > 0.05). In conclusion, HEPES and CO2 had no significant effect on swim-up of buffalo spermatozoa. The pH 8.5 of TALP improved the recovery rate of motile spermatozoa in swim-up procedure. The ion-exchange filtration was found superior to swim-up procedure in harvesting maximum number of motile spermatozoa from frozen-thawed buffalo semen (95 vs 33%; P < 0.001).

Animals↗
PubMed2001

Delayed suprachoroidal hemorrhage after glaucoma filtration procedures.

OBJECTIVES: To determine the incidence of, risk factors for, and outcomes of delayed suprachoroidal hemorrhage (DSCH) after glaucoma filtration surgery. DESIGN: Retrospective case-control study. PARTICIPANTS: All patients undergoing glaucoma filtration procedures between 1986 and 2000 at Indiana University who were diagnosed postoperatively with suprachoroidal hemorrhage. A total of 66 patients with DSCH were identified. These were compared with a randomly selected group of patients who underwent similar procedures but did not have suprachoroidal hemorrhage. METHODS: Total cases of DSCH were initially compared with the total number of glaucoma surgeries to determine the overall incidence and the incidence in the different procedures. Subsequently, a case-control study was performed comparing the group with hemorrhage to the control group to identify risk factors. Finally, outcomes and prognostic factors were determined by comparing vision preoperatively and postoperatively and parameters of patients with good and poor outcomes. MAIN OUTCOME MEASURES: Incidence of DSCH, risk factors associated with its occurrence, visual outcomes, and factors important for prognosis. RESULTS: Of a total of 2285 glaucoma filtration procedures, 66 (2.9%) cases of DSCH were identified. It developed in 9 of 615 (1.5%) trabeculectomies without antimetabolite, 30 of 1248 (2.4%) trabeculectomies with antimetabolite, 2 of 72 (2.8%) valved tube shunt implantations, and 25 of 350 (7.1%) nonvalved tube shunt implantations. The increased incidence of DSCH after tube shunts compared with trabeculectomy-associated DSCH was significant (P < 0.0001) with an odds ratio of 3.2. The risk factors for DSCH after glaucoma surgery include white race (P = 0.012), anticoagulation (P = 0.034), severe postoperative hypotony (P = 0.033), and aphakia/anterior chamber intraocular lens (P = 0.002). The visual outcomes of patients with hemorrhage were poor, with a decrease in logarithm of the minimum angle of resolution visual acuity from 0.72 to 1.36, which was statistically significant compared with the controls (P < 0.009). CONCLUSIONS: Delayed suprachoroidal hemorrhage occurs more frequently after tube shunt implantation than after trabeculectomy. Caution should be exercised when operating on patients with known risk factors, because the visual outcomes after DSCH are poor.

Aged↗
PubMed1999

Detection of Mycobacterium bovis lymphocyte stimulating antigens in culture filtrates of a recombinant Mycobacterium smegmatis cosmid library.

Culture filtrates derived from a Mycobacterium bovis cosmid library in Mycobacterium smegmatis were screened for bovine lymphocyte stimulatory antigens using peripheral blood mononuclear cells (PBMC) from cattle vaccinated with a low dose of Mycobacterium bovis BCG. Lymphocyte proliferation and interferon-gamma (IFN-gamma) production were used as cellular response markers for antigen recognition. In the primary screen, approximately 28% of all culture filtrates (CF) stimulated responses by PBMC from at least two out of four vaccinated cattle. In one of these CF, the M. bovis Ag85-B antigen was detected by Western-blot analysis. Despite heterogeneous lymphocyte responses of the animals, twenty-four of the culture filtrates stimulated lymphocyte proliferation and IFN-gamma production from at least six out of eight vaccinated animals in a secondary screen. Analysis of the cosmid DNA associated with these positive CF demonstrated that several contained homologous DNA sequences. It appears that the lymphocyte screening has detected M. bovis antigens that are immuno-dominant in cattle vaccinated with M. bovis BCG.

Adjuvants, Immunologic↗
PubMed1998

Isolation of the factor VIII-von Willebrand factor complex directly from plasma by gel filtration.

A high capacity gel filtration system was developed with the purpose of isolating factor VIII (FVIII) and von Willebrand factor (vWF) directly from plasma in significantly higher yields than obtained by cryoprecipitation, the technique most commonly used to recover FVIII-vWF from human plasma. After laboratory-scale gel filtration of plasma, a FVIII-containing fraction was collected containing about 90% of FVIII in the applied plasma and with almost tenfold higher purity than that obtained by cryoprecipitation. The gel filtration step has been scaled up for use as the initial step in the manufacturing process for a FVIII preparation (Nordiate).

Chromatography, Gel↗
PubMed1999

Cell salvage in obstetrics: an evaluation of the ability of cell salvage combined with leucocyte depletion filtration to remove amniotic fluid from operative blood loss at caesarean section.

During 27 elective caesarean sections, operative blood loss was collected and processed using the Haemonetics Cell Saver 5 and filtered by Pall RC 100 leucocyte depletion filtration. The efficiency of removal of amniotic fluid, and the degree. of contamination with fetal red cells were assessed in the resulting 'cleaned' blood. Cell saver processing effectively removed alpha-fetoprotein from the red cells of 14 patients whose amniotic fluid was removed by separate suction and from nine of the 13 patients whose amniotic fluid was aspirated into the cell saver along with operative blood loss. Cell saver processing and leucocyte depletion filtration completely removed trophoblastic tissue and white cells, but fetal squames were still clearly present in 10, and possibly in 14 samples after processing and fully removed in only two specimens. Amorphous debris was present in all samples after processing. The maximum mass of fetal red cells contaminating any patient's total salvaged blood was 19 ml (range 2-19 ml). Had this been re-transfused into a rhesus-incompatible mother it would have required 2500 i.u. (500 microg) anti-D immunoglobulin to prevent rhesus-immunization of the mother. Contamination of processed caesarean section blood with fetal red cells and fetal squames is defined and its clinical implications discussed, with an overview of the development and current status of cell salvage. Autotransfusion by cell salvage with leucocyte depletion filtration should be considered in life-threatening obstetric haemorrhage and offered to Jehovah's Witnesses.

Journal Article↗
PubMed2003

Replacement of maize by rumen filtrate fermented corn-cob in layer diets.

Laying hens (50 weeks in-lay) were fed for 8 weeks on diets containing corn-cobs fermented with rumen filtrate. The fermented corn-cob was included in the diets at 0%, 5.0%, 10.0%, 15.0% and 20.0% of the total diet at the expense of maize. Feed intake was not significantly (P>0.05) different, although birds on the control diet consumed slightly more feed than birds on the fermented corn-cob based diets. Hen day production and feed per dozen eggs decreased with increase in dietary level of corn-cob (P<0.05). Yolk colour was significantly (P<0.05) affected, rumen filtrate fermented corn-cob appear to promote better yolk coloration. Egg weight and shell thickness were not significantly influenced by dietary treatment. The overall results appear to suggest that rumen filtrate fermentated corn-cob could provide a material that holds promise as a good alternative to maize because of its enhanced crude protein value and reduced crude fibre content.

Animal Feed↗
PubMed2001

Mechanical thrombolysis of venous thrombosis in an animal model with use of temporary caval filtration.

PURPOSE: To test the effect of temporary caval filtration on pulmonary emboli when a mechanical thrombolytic device is used to treat venous thrombosis and to test the effects of a modified device on caval patency at 30-day follow-up. MATERIALS AND METHODS: In a canine model of iliocaval subacute thrombosis, mechanical thrombolysis was performed with use of an 8-F over-the-wire Arrow-Trerotola Percutaneous Thrombolytic Device (PTD) with a 9-mm (iliac) or 15-mm (inferior vena cava [IVC]) basket. In six procedures, the device was made of nitinol monofilament, and in another six, it was made of braided stainless steel. All procedures were performed with a nitinol expandable sheath (temporary filter) in the suprarenal IVC. Low-molecular-weight heparin was given daily after the procedure. Venography, pulmonary arteriography, measurement of blood gases, and pulmonary artery (PA) pressure measurement were performed before and after the procedure and at 30-day follow-up. Pulmonary arteriograms from the group treated with stainless-steel devices were compared to those from an earlier group of animals in which the identical procedure was performed without caval filtration. The IVC was examined histologically. RESULTS: Thrombolysis was successful in all animals. Rare segmental and subsegmental pulmonary emboli (PE) were seen arteriographically; compared to procedures without filters, there was a significant reduction in PE (P <.002). However, a mild increase in pulmonary artery pressure, decrease in pH, and increase in pCO(2) were observed postprocedurally. At 30-day follow-up (n = 11), IVC patency was preserved in 45% (n = 5) of animals overall. Caval patency was significantly better in animals in which the combination of stainless-steel devices was used (five of six = 83% vs zero with nitinol device; P =.015). Histologically, the stainless-steel device caused little intimal injury and fibrosis-less than that seen with the nitinol device. CONCLUSIONS: Temporary filtration reduces, but does not completely eliminate, PE during mechanical thrombolysis. The stainless-steel device results in less intimal injury and better caval patency than the nitinol device.

Animals↗
PubMed1976

Multiple forms of oviduct progesterone receptors analyzed by ion exchange filtration and gel electrophoresis.

Resolution of the multiple forms of steroid receptors in small samples has been improved by two new techniques: preparative ion exchange filtration and electrophoresis in highly cross-linked polyacrylamide gels of varied concentration. These techniques were used in conjunction with protamine precipitation, gel filtration, and density gradient centrifugation to separate five forms of the progesterone receptor of chick oviduct cytosol. These complexes, numbered I to V in order of elution from agarose gel columns, have been characterized with respect to apparent molecular weight, shape, and relative net charge. Form I, which is eluted in the void volume after gel filtration of cytosol in hypotonic media, is heterodisperse with respect to sedimentation coefficient and electrophoretic mobility (Rf). Form I is converted to form III by KC1. Form II has the highest axial ratio and the highest Rf at pH 10.2. This 4.2S complex can be extracted from DEAE filters, but not from protamine-precipitated cytosol, by 0.3 to 0.5 M KC1. Form III is slightly smaller (3.9S) and less asymmetric than form II. It is relased from DEAE filters and protamine-precipitated cytosol by 0.15 M KC1 and displays increased Rf upon purification. Forms II and III correspond to the B and A components described by W. T. Schrader and B. W. O'Malley ((1972), J. Biol. Chem 247, 51). Form IV may result from the proteolytic cleavage of forms II and/or III. Form V is a globular polypeptide obtained in the presence of certain divalent cations. This complex has been named the "mero-receptor" since it is the smallest part or fragment of the receptor that contains the steroid-binding site.

Animals↗
PubMedDate not supplied

A new integrated membrane filtration and chromatographic device.

To improve protein separation, a novel integrated device combining membrane filtration and chromatography has been developed. The device basically consists of a hollow fiber filtration module whose shell side is filled with chromatographic resin beads. However, there is an essentially impermeable coated zone near the hollow fiber module outlet. The integrated device enjoys the advantages of both membrane filtration and chromatography; it also allows one to load the chromatographic media directly from the fermentation broth or lysate and separate the adsorbed proteins through the subsequent elution step in a cyclic process. Interfacial polymerization was carried out to coat the bottom section of the hollow fiber membrane; the rest of the hollow fiber membrane remained unaffected. Myoglobin (Mb) and alpha-lactalbumin (alpha-LA) were primarily used as model proteins in a binary mixture; binary mixtures of Mb and bovine serum albumin (BSA) were also investigated. Separation behaviors of binary protein mixtures were studied in devices having either an ultrafiltration (UF) or a microfiltration (MF) membrane. Experimental results show that the breakthrough time and the protein loading capacities were dramatically improved after introducing the impermeable coating in both UF and MF modules. For a synthetic yeast fermentation broth feed, four loading-washing-elution-reequilibration-based cyclic runs for separation of Mb and alpha-LA were performed in the device using a MF membrane with a coated zone without cleaning in between. The Mb and alpha-LA elution profiles for the four consecutive runs were almost superimposable. Due to lower transmembrane flux in this device plus the periodical washing-elution during the chromatographic separation, fouling was not a problem, unlike in conventional microfiltration.

Chromatography, Liquid↗