Differences In Migration Timing Research Articles

Determination of Carbohydrate-deficient Transferrin Using Capillary Zone Electrophoresis

Current methods for carbohydrate-deficient transferrin (CDT) often suffer from low precision, complexity, or risk of false positives attributable to genetic variants. In this study, a new capillary zone electrophoresis (CZE) method for CDT was developed. CZE was performed on a P/ACE 5000 using fused-silica capillaries [50 microm (i.d.) x 47 cm] and the CEOFIX CDT buffer system with addition of 50 microL of anti-C3c and 10 microL of anti-hemoglobin. Native sera were loaded by high-pressure injection for 3 s, separated at 28 kV over 12 min, and monitored at 214 nm. CDT was completely resolved by differences in migration times (di-trisialotransferrin, 9.86 +/- 0.05 min; monosialotransferrin, 9.72 +/- 0.05 min; asialotransferrin, 9.52 +/- 0.04 min), with a CV of 0.15%. The number of theoretical plates was 312,000 +/- 21,000 for the mono- and 199 000 +/- 6500 for the di-trisialylated transferrin. Genetic CB and CD variants showed prominent peaks with migration times of 10.12 +/- 0.06 and 9.89 +/- 0.03 min, respectively, and the carbohydrate-deficient glycoprotein syndrome could be detected, excluding false-positive results. CZE results (as a percentage; y) correlated with the Axis %CDT TIA (x) values by Deming regression analysis: y = 1.92x - 7.29; r = 0.89. CDT values in 130 healthy nonalcoholics were determined. The 2.5th and 97.5th percentiles were 1.84% and 6.79%. CZE without sample pretreatment can determine CDT with good precision, allows detection of variants, and correlates with ion-exchange chromatography.

A new method for fast haptoglobin phenotyping and hemoglobin binding capacity calculation based on capillary zone electrophoresis.

A capillary zone electrophoresis method was developed for haptoglobin (Hp) phenotyping in hemoglobin (Hb) supplemented serum. The method allows a complete resolution of the major haptoglobin phenotypes Hp 1-1, Hp 2-1, and Hp 2-2 based on the difference in charge-to-mass ratio of their Hb-Hp complexes. Identification of these phenotypes was achieved by their significant differences in migration times and their marked difference in electrophoretic pattern. Our method showed full agreement with starch gel electrophoresis. Furthermore, following neuraminidase treatment of the serum, the Hp subtypes Hp1-1FF, FS and SS could be resolved, based on the same criteria as the phenotyping. The new electrophoretic method allowed typing of the rare phenotypes Hp 2-1 modified (Hp 2-1M) and Hp Johnson. The calculated hemoglobin binding capacity of serum correlates well with the nephelometrically determined haptoglobin concentration. The new method for typing haptoglobin gives prospectives for fast haptoglobin typing and Hp 1-1 subtyping.

Discrimination of granulocyte colony-stimulating factor isoforms by high-performance capillary electrophoresis

Granulocyte colony-stimulating factor (G-CSF) is a glycoprotein which acts primarily to stimulate the proliferation, differentiation and activation of committed progenitor cells of the neutrophil–granulocyte lineage into functionally mature neutrophils. The traditional biological assays employed to detect G-CSF are a myeloid bone marrow colony assay, a factor-dependent cell line specific for G-CSF and commercially available immunoassays. However, these methods will not distinguish between glycosylated and non-glycosylated forms of the molecule. In this study high-performance capillary electrophoresis (HPCE) was used to analyse glycosylated and non-glycosylated recombinant human granulocyte colony-stimulating factor (r-met-hG-CSF). Glycosylated G-CSF preparations contained human serum albumin (HSA), added as a protein carrier. Glycosylated and non-glycosylated G-CSFs were prepared in 40 m M Na 2HPO 4 buffer, pH 2.5, containing hydroxypropylmethylcellulose (HPMC) or 50 m M Na 2HPO 4 buffer, pH 9.0. Glycosylated G-CSF could be separated into two distinct glycoform populations at the lower pH studied. Differences in migration time and peak shape between glycosylated and non-glycosylated G-CSF were demonstrated. HPCE analysis of G-CSF produced using a baculovirus expression vector system revealed a further distinct G-CSF glycoform and demonstrated the resolving power of the technique.

Migration Timing of Atlantic Salmon Smolts Relative to Environmental and Physiological Factors

We determined the migration timing of fry-stocked smolts of Atlantic salmon Salmo salar relative to environmental and physiological factors, by using net weirs and counting fences in three tributaries of the West River, Vermont. Smolt migration began in late April and early May when water temperature was 5°C, peak movements occurred in early and mid-May at temperatures exceeding 8°C, and migration was complete by early June. Within this seasonal window, significant differences in migration timing and gill Na+,K+-ATPase activity occurred among tributaries. In both years of the study, smolts tended to migrate earlier and exhibit greater gill Na+,K+-ATPase activity in the warmest tributary than in the coolest tributary. Smolt migration timing differed most among tributaries in mid-May when (1) water temperatures were more than 8°C, (2) smolts peaked in gill Na+,K+-ATPase activity, and (3) discharge peaked, stimulating smolt migration. Smolts captured after the migratory period had lower gill Na+,K+-ATPase activity than migrating smolts. Relating smolt physiology to migration was crucial for explaining complex interactions among water temperature, discharge, and smolt behavior during both the onset and cessation of migratory activity. Because the period between onset of migration and loss of smolt physiological characteristics may be brief, delays in downstream passage that may occur at dams must be minimized to maximize the successful recruitment of smolts to the marine environment.

Capillary Zone Electrophoresis Separation of Amino Acid Enantiomers as Dansylated Derivatives Through Control of Electroosmotic Flow

D,L-Amino acid enantiomers are separated by capillary zone electrophoresis (CZE) as 5-dimethylamino- 1-naphthalene sulfonyl (dansyl) derivatives using a Cu2+-L-aspartyl-L-phenylalanine methyl ester (aspartame) complex as a chiral selector in the buffer. Only partial enantiomeric resolution is obtained for several dansyl-D,L-amino acids and for the most favorable cases resolution approaches about 1.4. Increasing Mg2+, Cd2+, or Zn2+ concentration in a 10 mM NH4OAc, 2.5 mM Cu2+, 5.0 mM aspartame, pH = 7.40 buffer increases dansyl-D,L-amino acid migration time, increases migration time difference between the D-enantiomer, which appears first in the electropherogram, and the L-enantiomer, and resolves the enantiomers of most dansyl-D,L-amino acids with resolution values in the range of 1.4 to > 5.0 depending on the derivative and buffer conditions. An increase in the buffer divalent cation concentration reduces the electroosmotic flow (EOF) while the electrophoretic mobilities of the dansyl-D,L-amino acid enantiomers do not undergo significant change as the divalent cation concentration increases. The improved resolution between the dansyl-D,L-amino acid enantiomers is due to the reduced EOF and small differences in the ternary complex formed between the dansyl-D,L-amino acid enantiomers and the Cu2+-aspartame complex. †nee, B. Chanthawat.

Electrokinetic chromatography in suppressed electroosmotic flow environment: Use of a charged cyclodextrin for the separation of enantiomers and geometric isomers

Electrokinetic chromatography (EKC), with negatively-charged cyclodextrins (NCDs) added to the buffer, was conducted in polyacrylamide-coated columns under suppression of electroosmotic flow. The equations of migration and resolution for neutral solutes in this mode of chromatography, which for brevity we term NCD-EKC, are presented. The chiral sulfated cyclodextrin, beta-CD-SBE (IV), used in this study is anionic over the entire pH range accessible to capillary electrophoresis, and the coated columns are stable and provide reproducible performance in the pH range 2.5-8.8. Optimum separation was obtained in the pH range where the solutes are neutral. The incorporation of an alkyl spacer between the sulfate ion and the rim of the cyclodextrin allows an unhindered approach and inclusion of neutral solutes in the cyclodextrin cavity. Solute migration time is inversely proportional to the concentration of the chiral selector. Separation (relative migration time difference) increases with decreasing chiral selector concentration and approaches a maximum, beyond which further decreases in chiral selector concentration result in broad peaks and loss of resolution. A chiral selector concentration of 1% in a 10 mM phosphate buffer produced excellent separation of amino acids and dipeptide enantiomers. In addition to being chiral selectors, cyclodextrins are also known as shape selectors. NCD-EKC is particularly suited for the separation of positional isomers of hydrophobic solutes. The separation of aflatoxin isomers and chlorophenol congeners is presented. In the separation of chlorophenols the more hydrophobic trichlorophenols eluted first and the least hydrophobic, phenol, eluted last.

Separation of 8-aminonapthalene-1,3,6-trisulfonic acid-labelled neutral and sialylated N-linked complex oligosaccharides by capillary electrophoresis

Complex oligosaccharides, both neutral and sialylated, were derivatized with 8-aminonaphthalene-1,3,6-trisulfonic acid (ANTS) and separated by capillary electrophoresis. The derivatization reaction was carried out in a total reaction volume of 2 μl. The separated peaks were detected by laser induced fluorescence detection using the 325-nm line of a HeCd laser. Concentration and mass detection limits of 5·10 −8 M and 500 amol, respectively, could be achieved. The limiting step for higher sensitivity is not the detector performance, however, but the chemistry with a derivatization limit of 2.5·10 −6 M. Two labelling protocols were established, one with overnight reaction at 40°C and the other with a 2.5-h derivatization time at 80°C. Neutral oligosaccharides could be labelled with either protocol. However, sialylated oligosaccharides hydrolysed when labelled at 80°C. Low nanomole to with either protocol. However, sialylated oligosaccharides hydrolysed when labelled at 80°C. Low nanomole to picomole amounts of oligomannose-type and complex-type oligosaccharide mixtures were derivatized and separated in less than 8 min with excellent resolution using a phosphate background electrolyte at pH 2.5. The linear relationship between the electrophoretic mobility and the charge-to-mass ratios of the ANTS conjugates was used for peak assignment. Further, the influence of the three-dimensional structure of the complex oligosaccharides on their migration behaviour is discussed. The suitability of the ANTS derivatization and the subsequent separation for the analysis of complex oligosaccharide patterns is demonstrated with oligosaccharide libraries derived from ovalbumin and bovine fetuin. For peak assignment the patterns are compared with those of the oligomannose and the complex-type oligosaccharide mixtures. The separation efficiency of 120 000 theoretical plates and analysis times of less than 10 min are superior to those with state-of-the-art chromatographic methods and other capillary electrophoresis separation methods. A migration time difference of 0.06 min was found to be sufficient for the baseline separation of complex oligosaccharides.

Capillary gel electrophoresis of oligonucleotides: prediction of migration times using base- specific migration coefficients

Chemically synthesized oligodeoxyribonucleotides were subjected to capillary gel electrophoresis on three different polyacrylamide-based matrices. Analysis of about 1000 samples over a 1-year period showed that the gel matrix evolved with time resulting in shifting migration times, making it essential to use an internal standard. Cross-linked polyacrylamide matrices had the highest stability, allowing an average of 100 injections on the same capillary. Computer- aided prediction of migration times was subsequently evaluated to confirm the size and base composition of oligonucleotides more accurately. A number of problems were noted when using this approach on a routine basis, such as insufficient stability of the gel matrices, effects of secondary structure on migration and insufficient differences in migration times for oligonucleotides containing >50 bases. Capillary gel electrophoresis at pH 3.5 in replaceable gels showed that migration was mainly dependent on the charge per base ratio resulting in separations of significantly altered selectivity which complemented analyses under the commonly used basic pH conditions.

Differential Timing of Autumn Migration between Sex and Age Groups in Raptors at Falsterbo, Sweden

The migration of raptors at Falsterbo, Sweden was studied during five autumns, 1986-1990, to investigate the intraspecific temporal sequence of migration for different age and sex classes. The analysis is based on counts of between 45 and 72 321 individuals of 14 species. The percentage of juveniles varied among species and to some extent between years. This reflects a variable tendency of different migrants to concentrate at Falsterbo and fluctuating breeding success from year to year. Most species showed an even sex ratio. Marked exceptions were Marsh Harrier Circus aeruginosus and Hen Harrier C. cyaneus in which females constituted 60 and 61% respectively of adult migrants. This biassed sex ratio agrees with the tendency towards polygyny in these species. In tropical (long-distance) migrants adults migrated ahead of juveniles (Honey Buzzard Pernis apivorus, Montagu's Harrier Circus pygargus, Osprey Pandion haliaetus, and Hobby Falco subbuteo). It is suggested that it is advantageous for adults to arrive early on the wintering grounds, finish moult and accumulate energy reserves before spring migration and the next breeding season. Those short-distance migrants in which adults migrated before juveniles at Falsterbo are species whose majority leave Scandinavia in the winter (Hen Harrier, Common Buzzard Buteo buteo, Roughlegged Buzzard B. lagopus, Merlin Falco columbarius and Peregrine F peregrinus). Adults leaving first have a greater possibility to secure a good winter territory forcing the juveniles, migrating later, to settle in less suitable habitat or to winter further south. In species where juveniles migrated ahead of adults a large proportion of the adults spend the winter close to the breeding grounds (Red Kite Milvus milvus, Goshawk Accipiter gentilis and Sparrowhawk A. nisus). The presumably subdominant juveniles may be excluded from the breeding territories by the adults and therefore migrate south, followed by a varying proportion of adults. Females generally migrated before males at Falsterbo due to different moult strategies; females starting and finishing moult of the flight feathers before males. This difference in timing of moult and migration is ultimately dependent on a marked role division between the sexes during the breeding cycle. In Honey Buzzard where both sexes share the breeding duties equally no difference in timing of migration was found. Social dominance, importance of early arrival at the winter quarters and moult strategies are the factors best explaining the observed differential timing in migrating raptors at Falsterbo.

Bacterial migration along solid surfaces.

An in vitro system was developed to study the migration of uropathogenic Escherichia coli strains. In this system an aqueous agar gel is placed against a solid surface, allowing the bacteria to migrate along the gel/solid surface interface. Bacterial strains as well as solid surfaces were characterized by means of water contact angle and zeta potential measurements. When glass was used as the solid surface, significantly different migration times for the strains investigated were observed. Relationships among the observed migration times of six strains, their contact angles, and their zeta potentials were found. Relatively hydrophobic strains exhibited migration times shorter than those of hydrophilic strains. For highly negatively charged strains shorter migration times were found than were found for less negatively charged strains. When the fastest-migrating strain with respect to glass was allowed to migrate along solid surfaces differing in hydrophobicity and charge, no differences in migration times were found. Our findings indicate that strategies to prevent catheter-associated bacteriuria should be based on inhibition of bacterial growth rather than on modifying the physicochemical character of the catheter surface.

Life History and Timing of Migrations and Spawning Behavior of Rainbow Trout (Salmo gairdneri) Populations of the Great Lakes

Rainbow trout (Salmo gairdneri) populations of the Great Lakes showed a great variability in timing of spawning migrations and life histories. This variability was examined to determine if rainbow trout populations of the Great Lakes are comprised of discrete stocks. Differences in timing of migration and spawning indicated that at least one spawning population may be distinctive and that others may be in progressive stages towards emergence of discrete stocks. The innate ability of this species to adapt to different environmental conditions, together with its ability to 'home,' provide rainbow trout in the Great Lakes with the potential to develop discrete stocks. The management of this species that allows for the development of this potential is emphasized.Key words: rainbow trout. Great Lakes, stocks, migrations, spawning, life histories, homing