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PubMed1991

[Stem cell separation with different cell separators].

For some years, there has been an increasing success in transplanting peripheral blood stem cells (PBSC) instead of autologous bone marrow in patients suffering from different malignancies. While collecting PBSC for autologous transplantation, we compared four different separation techniques and three different cell separators (COBE Spectra, Fresenius AS 104, Haemonetics V50) routinely used for platelet production. Our results suggest that continuous flow separators seem to have some advantage over discontinuous flow machines in harvesting PBSC.

Blood Cell Count↗
PubMed1995

Preparation and storage in Plasma-Lyte A of platelets collected with the cell separator CS3000 Plus equipped with the PLT30-separation and TNX6 collection chambers.

The last five years have been characterized by the presentation of new cell separators the main task of which is the collection of high yield-high quality platelets. The CS3000 is an old apparatus which has undergone rejuvenation to sustain the assault of its new competitors. The CS3000 Plus Omnix system is the latest version to be offered along with a combination of TNX-6 separation/PLT30 collection chambers for optimal platelet collection. In this paper we present our results with this apparatus and configuration after its adaptation to the collection and storage of platelets in a non-plasma medium, the Baxter Plasma-Lyte A. After separation the platelet product (PC) was left in the collection chamber and resuspended with 200 ml of Plasma-Lyte A instead of being resuspended in autologous plasma as usual. Plasma was collected in a separate bag (400-450 ml) for transfusion or fractionation purposes. PC quality was assessed by evaluating the platelet yield (4.17 +/- 1.8 x 10(11), and the WBC contamination (4.8 +/- 2.6 x 10(5)). The presence of platelet aggregates (platelet count after aggregate fixation with formalin/platelet count after disaggregation in EDTA), the aggregation induced by ADP, collagen and ristocetin, the hypotonic shock response and the stability of membrane glycoproteins (CD 62 - 62 - 63 - 36- 42b - 51) were measured in the preapheresis samples and in the PC immediately after, 24 and 72 hours after collection. These results were totally satisfactory as was the post-transfusion survival measured as corrected count increment in 10 transfusions to non refractory patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1994

Responses of free-ranging rhesus monkeys to a natural form of social separation. I. Parallels with mother-infant separation in captivity.

Observations of 23 free-ranging rhesus monkey infants on Cayo Santiago, Puerto Rico, indicated that mothers' first postpartum estrous periods were marked by large increases in the amount of time infants were separated from their mothers, by disturbances in mother-infant relationships, and by increases in infant distress behavior. When their mothers resumed mating, most infants showed signs of agitation; a few briefly showed indications of depression. Male infants responded to their mothers' resumption of mating by playing more, whereas females engaged in less play and more allogrooming. The results suggest (a) that basic parallels exist between the behavioral responses of rhesus infants to their mothers' resumption of mating in the field and to forcible separation from their mothers in captivity and (b) that early separation experiences may play a role in the normal development or manifestation of sex differences in behavior.

Animals↗
PubMed2001

Protein from pea mutants as a co-product in starch separation--isolates from wet and dry separation: yield, composition and solubility.

The dry separation behaviour of cotyledon material of certain pea genotypes (wild type and 5 r mutants) with different metabolic background in starch biosynthesis allowed satisfying protein recovery only with smooth seeded genotypes. Structural effects and protein distribution (starch granule size distribution, protein in non-starch part) play an important role. According to expectations protein composition remained unaltered during dry separation procedures. Independent of genotypes used the applied conditions in wet separation allowed to recover just 50% of seed proteins while approximately 10% got lost with fibres and 40% with unspecified process water streams. Globular vicilins remained dissolved during iso-electric precipitation and were discarded with waste water streams. The solubility of isolates was rather low under alkaline conditions, even for the round seeded cv. Odalett (0.4 to 0.7% dry substance).

Genotype↗
PubMed2002

Size-based separation of synthetic polyelectrolytes in entangled polymer solution capillary electrophoresis: the effect of binary mixtures of separating polymers differing in molecular mass.

The influence on the electrophoretic behavior of polystyrenesulfonates of the percentage of high-molecular-mass chains in an entangled poly(ethylene oxide) solution having a bimodal molecular mass distribution has been investigated and compared with the results obtained for similar solutions of unimodal molecular mass distribution. The comparisons between the different separating polymer solutions were made at a constant total mass concentration, so as to keep constant the mesh size and to highlight the sole effect of the network dynamics. The use of binary polymer mixtures of two different molecular masses but of same nature can be a convenient alternative to modulate the dynamics of the network and the viscosity of the separating medium. A 20-30% content of high-molecular-mass chains in an entangled poly(ethylene oxide) solution having a binary molecular mass distribution appears to be a good compromise for a moderate viscosity and a good separation selectivity in comparison with a solution containing only chains of high molecular mass at the same concentration.

Electrophoresis, Capillary↗
PubMed2004

Matrix-assisted laser desorption/ionization-time of flight-mass spectrometry of lipopolysaccharide species separated by slab-polyacrylamide gel electrophoresis: high-resolution separation and molecular weight determination of lipooligosaccharides from Vibrio fischeri strain HMK.

We recently demonstrated that the combined use of lipopolysaccharide (LPS) reverse staining and high-efficiency passive elution techniques can be successfully used as a suitable interface between LPS slab-gel separation and electrospray ionization-mass spectrometry (ESI-MS) of LPS-derived oligosaccharides. Here, we extend our micropurification strategy for the analysis of O-deacylated LPS forms from Vibrio fischeri HMK after recovery from single reverse-stained LPS bands using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The quantities (30-40 microg) obtained from the two gel-resolved LPS bands were sufficient to allow MALDI-TOF-MS detection of O-deacylated LPS glycoforms at m/z 3767.1, 3890.1 for the high-molecular-weight or at m/z 2522.5, 2645.4, 2725.7, and 2848.7 for the low-molecular-weight LPS band. These LPS band heterogeneities resulted not only from variations in the oligosaccharide region of the LPS but also from two phosphorylation states of the lipid A (diphosphoryl and diphosphoryl plus a single phosphoethanolamine substitution). On the other hand, MALDI-TOF mass spectra of the separated LPS bands displayed reduced heterogeneity and increased signal-to-noise ratios as compared to spectra of the unpurified LPS. Furthermore, micropurification of LPS bands prior MALDI-TOF-MS led to a higher sensitivity of detection of less abundant low-molecular-weight LPS glycoforms. Taken together, this and our previous study on gel-micropurified LPS using ESI definitively show how one can unambiguously determine the different molecular species contained within each gel-separated LPS band, their relative abundance and oligosaccharide sequences.

Aliivibrio fischeri↗
PubMed1993

Novel affinity separations based on perfluorocarbon emulsions. Development of a perfluorocarbon emulsion reactor for continuous affinity separations and its application in the purification of human serum albumin from blood plasma.

Perfluorocarbon affinity emulsions are generated by the homogenisation of a perfluorocarbon oil with a polymeric fluorosurfactant previously derivatised with an affinity ligand and subsequently cross-linked in situ. This procedure gives rise to a novel liquid affinity adsorbent that can be used for continuous protein purification. Discrete emulsion droplets were found to be unstable when pumped for prolonged periods; however, when flocculated, the emulsion floccules with diameters of around 125 microns, were very stable and sedimented faster. A four-stage reactor unit (perfluorocarbon emulsion reactor for continuous affinity separations, PERCAS) was designed and constructed to carry out continuous separations, and exploited the unusual properties of the absorbent, i.e. liquid nature and high density. Each of the four stages of PERCAS consisted of a mixing tank, for contacting between emulsion phase and aqueous phase, adjacent to a settling tank for the subsequent separation of emulsion from the aqueous phase. Using PERCAS adsorption, washing, elution and re-equilibration of the emulsion could be carried out continuously with emulsion recycle. Using single-component adsorption of human serum albumin to a perfluorocarbon affinity emulsion derivatised with the triazine dye C.I. Reactive Blue 2, PERCAS was optimised with respect to flow-rates and input concentrations. The work was then extended to the continuous purification of essentially homogeneous human serum albumin from blood plasma.

Adsorption↗
PubMed1994

Temperature as a variable in reversed-phase high-performance liquid chromatographic separations of peptide and protein samples. I. Optimizing the separation of a growth hormone tryptic digest.

Peptide and protein samples are often complex mixtures that contain a number of individual compounds. The initial HPLC separation of such samples typically results in the poor resolution of one or more band pairs. Various means have been suggested for varying separation selectivity so as to minimize this problem. In this study of a tryptic digest of recombinant human growth hormone, the simultaneous variation of temperature and gradient steepness was found to be a convenient and effective means of varying selectivity and optimizing the separation. The use of computer simulation greatly facilitated this investigation.

Chromatography, High Pressure Liquid↗
PubMed1989

Sequence of centromere separation: kinetochore formation and DNA replication in dicentric chromosomes showing premature centromere separation in rat cerebral cells.

A subpopulation of rat cerebral endothelial cells, designated B1, exhibits an array of multicentric chromosomes. Because of the formation of bridges at anaphase, this cell population produced new types of multicentrics at every cell division. These chromosomes showed kinetochore proteins at every centromeric site and all centromeric regions replicated their DNA at the end of the S phase, more or less simultaneously. A new subpopulation of cells, designated B2, obtained from the original sample frozen at Wayne State University displayed several dicentrics. In contrast to B1 these chromosomes exhibit premature centromere separation as reported for mouse and human cell lines. These B2 dicentrics show only one site of kinetochore protein deposition. The timing of DNA replication around the centric region of prematurely separating centromere is also changed similar to the earlier reported premature DNA synthesis for mouse dicentrics. These observations suggest a universality of relationship between premature centromere separation, a lack of kinetochore formation, and early replication of the centric/pericentric DNA associated with these centromeres. The cause of sudden change from activity to inactivity of these chromosomes, though interesting, is not clear.

Animals↗
PubMed2005

Comparison of a static gravity screen-roll press combination separator to a PAM-assisted gravity belt thickener system for swine waste slurry solids separation.

Liquid effluent resulting from solids separation from raw swine slurry (RS) using a static gravity screen-roll press separator (SE) and a polyacrylamide (PAM) flocculant-assisted gravity belt thickener system (BE1 and 2, with BE2 designating an added belt thickening of SE) were compared. Effluent pH was 7.6-7.7. Dissolved oxygen was less than 0.01 mg/l in RS and SE and 0.50 mg/l in BE1 and 2. Solids dry weight and total phosphorus concentrations were reduced significantly (p0.05) in SE (17-27%) and in BE1 and 2 (92-96%), relative to RS. Phosphorus concentration reductions were higher than those previously found. Settleable and suspended solids, total nitrogen and chemical oxygen demand were significantly reduced in BE1 and 2 (63-99%). No significant differences (p>0.05) were found between BE1 and 2. Results indicated that the gravity belt thickener system was more efficient at solids separation than the screen-roll press system.

Acrylic Resins↗
PubMed1985

Comparison of reversed-phase and cation-exchange high-performance liquid chromatography for separating closely related peptides: separation of Asp76-human parathyroid hormone (1-84) from Asn76-human parathyroid hormone (1-84).

Cation-exchange high-performance liquid chromatography (CE-HPLC) was compared with ordinary reversed-phase high-performance liquid chromatography (RP-HPLC) for separating closely related peptides. Some synthetic samples of bradykinin and angiotensins, which were homogeneous according to RP-HPLC, were found to be inhomogeneous when analyzed by CE-HPLC. On the other hand, diastereomeric peptides could be separated much more efficiently by RP-HPLC than by CE-HPLC. These results indicated that the purity of synthetic peptides should be checked not only by RP-HPLC but also by ion-exchange HPLC. In the case of human parathyroid hormone (hPTH), baseline separation of Asp76-hPTH from Asn76-hPTH by RP-HPLC was not possible, but was by CE-HPLC. Using this method we confirmed that the Asn residue in hPTH at position 76 could not be converted into the Asp residue under the conditions used to isolate and purify it from human organs.

Angiotensin II↗
PubMed1994

Temperature as a variable in reversed-phase high-performance liquid chromatographic separations of peptide and protein samples. II. Selectivity effects observed in the separation of several peptide and protein mixtures.

Changes in band spacing as a function of temperature and/or gradient steepness were investigated for four peptide or protein samples. Reversed-phase HPLC in a gradient mode was used to separate tryptic digests of tissue plasminogen activator and calmodulin. Additionally, a synthetic peptide mixture and a storage protein sample from wheat were studied. Simultaneous changes in gradient steepness and temperature were found to provide considerable control over band spacing and sample resolution. The effects of temperature and gradient steepness on selectivity in these systems appear to be complementary. Simultaneous optimization of both temperature and gradient steepness thus represents a powerful and convenient means of controlling band spacing and separation. Because of the complexity of these sample chromatograms, computer simulation proved to be a useful tool in both interpreting these experiments and in optimizing final separations.

Calmodulin↗
PubMed1996

Separation of chiral polychlorinated biphenyls by micellar electrokinetic chromatography using beta- and gamma-cyclodextrin mixtures in the separation buffer.

Chiral polychlorinated biphenyls (PCBs) 45, 84, 88, 91, 95, 132, 136, 139, 149, 171, 183 and 196 were separated each in its two enantiomers by cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC). Mixtures of beta- and gamma-cyclodextrins were used as chiral modifiers in a 2-(N-cyclohexylamino)ethanesulphonic acid (CHES) buffer containing urea and sodium dodecyl sulphate (SDS) micelles. Separations of multicomponent mixtures of PCBs into their enantiomers were also performed. A mixture of PCBs 45, 88, 91, 95, 136, 139, 149 and 196 was separated into all sixteen enantiomers in an analysis time of approx. 35 min.

Chromatography, Affinity↗
PubMed1999

Effect of electric field on liquid chromatographic separation of peptide digests. Combining capillary separation techniques.

A system is described which allows operation of a range of capillary based liquid phase separations including capillary electrophoresis, isocratic and gradient capillary electrochromatography, isocratic and gradient capillary liquid chromatography and electrically assisted gradient capillary liquid chromatography. The system was coupled to electrospray ionization mass spectrometry in the electrically assisted capillary liquid chromatography mode to investigate the effect of applied voltage on the selectivity in peptide mapping separations. Analyses were performed on tryptic digests of recombinant human growth hormone and tissue plasminogen activator. The results show a small but useful effect on selectivity that can be used to fine tune specific separations.

Algorithms↗
PubMed2001

Separation of DNA fragments and single strand conformation polymorphism analysis in bare capillaries using poly(acrylamide-dimethylacrylamide) as a separation medium.

A short chain poly(acrylamide-dimethylacrylamide) (PADMA) was synthesized in aqueous phase using isopropanol as a chain transfer agent, and was characterized according to the chemical composition and molecular mass. This polymer can form a stable dynamic coating on the inner surface of the capillary, thereby suppressing the electroosmotic flow and DNA-capillary wall interaction. The sieving medium has low viscosity and capillary filling with this medium and medium replacement were conveniently carried out by commercial capillary electrophoresis instruments. The effects of components and concentration of copolymers on the separation of DNA fragments were investigated. Highly efficient separation of DNA fragments, successful single strand conformation polymorphism (SSCP) analysis and good reproducibility of the migration time were obtained in bare capillaries using these copolymers as sieving media. Our preliminary results demonstrate that PADMA will become an alternative matrix for DNA separation by capillary electrophoresis.

Acrylamides↗
PubMed1998

Molded rigid polymer monoliths as separation media for capillary electrochromatography. 2. Effect of chromatographic conditions on the separation.

The effect of chromatographic conditions on the performance of monolithic poly(butyl methacrylate-co-ethylene dimethacrylate-co-2-acrylamido-2-methyl-1-propanesulfonic acid) columns in capillary electrochromatography has been studied. The flow velocity was found to be proportional to the strength of the electric field and both the pH and the composition of the mobile phase. A column efficiency of 120,000 plates/m at the optimum flow velocity of 1.5 cm/min is achieved for all the monolithic capillary columns of identical composition and porosity, regardless of their length, which varied from 30 to 120 cm. The polymeric separation medium exhibits retention and selectivity properties similar to those of typical ODS packings for reversed-phase chromatography. In addition to the "classical" use of monolithic capillary columns for the electrochromatographic separation of small molecules in reversed-phase mode, larger styrene oligomers were also separated under isocratic elution conditions. In addition, the electroosmotically driven size exclusion chromatography of polystyrene standards with molecular weights up to 10(6) has been demonstrated for the first time.

Benzene Derivatives↗
PubMed1979

The separation of peptide hormone diastereoisomers by reverse phase high pressure liquid chromatography. Factors affecting separation of oxytocin and its diastereoisomers--structural implications.

Experimental conditions and parameters involved in high performance liquid chromatography (HPLC) separations of the peptide hormone oxytocin and seven of its diastereoisomers, namely [1-hemi-D-cystine]-, [2-D-tyrosine]-, [4-D-glutamine]-, [5-D-asparagine]-, [6-hemi-D-cystine-], [7-D-proline]-, and [8-D-leucine]-oxytocin, on reverse phase columns were investigated. The effects of solvent, pH, and salt concentration were studied. Using the solvent systems 10% tetrahydrofuran-ammonium acetate buffer or 18% acetonitrile-ammonium acetate buffer and the muBondapak C18 support, oxytocin was separated from each of its diastereoisomers under all conditions studied, but the order of elution of diastereoisomers was highly dependent on solvent and to a lesser extent on pH. Separations of the hormone and its diastereoisomers on reverse phase HPLC and on classical partition chromatography on Sephadex G-25 were compared. The results are discussed in terms of the interactions of the solute with the reverse phase column and the solvent system. Implications of these findings in terms of the different solution conformations of the peptides are discussed.

Amino Acid Sequence↗
PubMed1992

Separation-individuation, family functioning, and psychological adjustment in college students: a construct validity study of the Separation-Individuation Test of Adolescence.

We evaluated the construct validity of the Separation-Individuation Test of Adolescence (SITA; Levine, Green, & Millon, 1986). Based on a review of the theoretical literature on separation-individuation issues during late adolescence, predictions were made as to how SITA scores would correlate with aspects of family functioning and psychological adjustment. The SITA and a battery of self-report family and adjustment measures were administered to 454 college students (272 females and 182 males; 74% White, 16% African American, and 10% Asian American or Hispanic). Correlational analyses offered partial support for the construct validity of the SITA. Cluster analyses revealed four distinct groups of subjects (anxious deniers, healthy separators, peaceful detachers, and succorance seekers), each with its own coherent set of scores on the SITA and instruments measuring family relations and positive and negative psychological adjustment. Criticisms concerning the labels and composition of the SITA scales are also discussed.

Adaptation, Psychological↗