Integrating microfabricated fluidic systems and NMR spectroscopy

The philosophy of miniature total analysis systems (mu-TAS) hinges on the integration of multiple chemical processing steps and the means of analyzing their results on the same miniaturized system. We have constructed chip-based capillary electrophoresis (CE) devices equipped with an integrated planar radio-frequency detector coil used for nuclear magnetic resonance spectroscopy (NMR). Separations were accomplished in the devices, but satisfactory NMR spectra could only be obtained from samples of high concentration. The relative sensitivity is explained and the scaling law dichotomy of CE and NMR explored.     

Effect of dietary IgG source (colostrum, serum, or milk-derived supplement) on the efficiency of Ig absorption in newborn Holstein calves

This study was designed to compare the absorptive efficiency of IgG from a commercial bovine serum product (bovine serum), cow colostrum (positive control), and two commercial milk-derived IgG supplements (supplement 1 and supplement 2). Newborn Holstein calves, collected at birth and prior to the consumption of colostrum, were allotted to treatment by alternating birth order. Colostrum supplement treatments were fed according to manufacturer's recommendations at birth and again at 12 h. This strategy resulted in varying masses of total IgG being offered to the calves (200, 90, 50, and 60 g of IgG for colostrum, bovine serum, supplement 1, and supplement 2, respectively). Blood samples were collected at 0, 12, and 24 h after the end of treatment administration. Plasma volume was estimated as 9.10% of birth weight. Apparent efficiency of IgG absorption at 24 h was determined. Plasma IgG concentrations at 24 h differed for each treatment (12.1, 6.8, 2.2, and 3.5 g of IgG/L for colostrum, bovine serum, supplement 1, and supplement 2, respectively). Apparent efficiency of IgG absorption was greatest for bovine serum compared with colostrum and supplement 1. No treatment differences were detected on the occurrence of mortality. However, calves fed bovine serum tended to have fewer treatments for illness compared with calves fed colostrum and supplement 1. Calves receiving bovine serum-derived IgG had improved IgG absorption efficiency and a tendency toward fewer medical treatments compared with calves consuming colostrum or a dried colostrum product.     

Passive immunoglobin transfer in newborn calves fed colostrum or spray-dried serum protein alone or as a supplement to colostrum of varying quality

Two experiments were conducted to investigate the effect of serum-derived immunoglobin (Ig) source and the effect of colostrum supplementation with serum-derived Ig on the attainment of passive immunity in newborn colostrum-deprived calves. In experiment 1, colostrum-deprived Holstein bull calves were fed pooled colostrum (PC, n = 9), spray-dried bovine serum (BS, n = 11), or spray- dried porcine serum (PS, n = 9). All treatments were balanced to provide 45 g of IgG in a 2-L volume at birth and again 12 h later. Calves receiving BS had higher 24-h serum IgG concentrations than did calves receiving PC or PS (8.3, 5.7, and 4.2 g of IgG/L for BS, PC, and PS, respectively). In experiment 2, the effect of supplementing bovine colostrum of varying quality with BS on Ig absorption was assessed. Thirty-two colostrum-deprived Holstein bull calves and four freemartin heifer calves were allotted by birth order to receive one of three treatments. Treatments consisted of 1) 2 L of pooled high quality colostrum (95.8 g of IgG, 0% from BS), 2) 2 L of pooled medium quality colostrum mixed with BS (95.2 g of IgG, 47% from BS), or 3) 2 L of low quality colostrum mixed with BS (98.8 g of IgG, 70% from BS). Serum IgG concentrations at 24 h after treatment were greater for calves receiving medium and low quality colostrum supplemented with BS (6.2, 9.6, and 9.6 g of IgG/L for high, medium, and low quality colostrum, respectively). Similarly, apparent efficiency of IgG absorption was greater for calves receiving medium and low quality colostrum supplemented with BS (25, 37, and 38% for high, medium, and low quality colostrum, respectively). The results of these studies suggest that dried BS contains a concentrated source of Ig, which is efficiently absorbed by newborn calves. Supplementation of marginal or low quality colostrum with dried BS is an effective means of improving passive transfer of IgG in newborn calves.     

Feeding linseed to increase the n-3 PUFA of pork: fatty acid composition of muscle, adipose tissue, liver and sausages

Eighty pigs, male and female littermate pairs, were fed a control or a test diet from 25 to 95 kg live weight. The diets, as fed, contained 15.5 g/kg linoleic acid (18:2) and 1.9 g/kg α-linolenic acid (18:3) (control) or 10 g/kg linoleic acid and 4 g/kg α-linolenic acid (test). The test diet, with added linseed, was, therefore, high in the main n-3 polyunsaturated fatty acid (PUFA) 18:3 and low in the main n-6 PUFA 18:2. Making this relatively small change led to a 56% increase in the content of 18:3 in muscle and major increases in the contents of the beneficial longer chain PUFAs EPA (20:5n-3) (100% increase) and DHA (22:6n-3) (35% increase) which are synthesised from 18:3n-3. Levels of EPA and DHA in pigmeat adipose tissue were also increased by the test diet. In liver, the test diet resulted in an 18:3 level 4× higher than in muscle, with 10× more EPA and 20× more DHA. Sausages, analysed after 6 months frozen storage also had high n-3 PUFA levels, due to the contribution of these fatty acids from both muscle and adipose tissue. From a health perspective these results confirm the potential of pigmeat to supply valuable n-3 PUFA to the human diet. The test diet produced a PUFA:saturated FA ratio in muscle of 0.4, close to the minimum recommended value for the diet as a whole and an n-6:n-3 ratio of 5, a significant improvement on the current average for pigmeat (7). It is estimated that the test diet would provide 12 g of long chain n-3 PUFA to the human diet per annum at current pigmeat consumption levels in the UK, about a third of that from oily fish.     

Sire variation in fatty acid composition of crossbred Wagyu steers and heifers

Effects of sires on lipid composition of subcutaneous adipose tissue and longissimus dorsi muscle were studied using 113 F(1) heifer and steer calves sired by eight Wagyu bulls out of three different cow herds. Wagyu sires were used and grouped as Old (n=6) and New (n =2) sires, respectively, based on the chronological order in which they were imported into the US. Animals were fed a backgrounding diet for 112 days consisting of an 80:20 ratio of roughage:concentrate, then grazed on orchard grass and bluegrass for 84 days, and finished on a 10:90 ratio of roughage:concentrate diet for 231 days in a feedlot. For longissimus dorsi muscle, progeny from Old sires had higher (P<0.05) monounsaturated to saturated fatty acid ratios (MUFA:SFA) than progeny of New sires. There were also differences (P<0.05) among individual sires for polyunsaturated to saturated fatty acid ratio (PUFA:SFA) (0.05-0.08) and MUFA:SFA (1.03-1.21). Progeny of Angus cows at Washington State University (WSU) had lower (P<0.05) MUFA:SFA and lower SFA than progeny of WSU crossbred and commercial cows. Steers had lower (P<0.05) MUFA:SFA and higher (P<0.05) SFA than heifers. For subcutaneous fat, heifers had higher levels (P<0.05) of linoleic acid (C18:2) and PUFA:SFA than the steers. Means for ether extractable fat in longissimus dorsi muscle differed among sires (P<0.05) and ranged from 7.58 to 13.13%. Progeny from WSU Angus cows had higher (P<0.05) ether extractable fat than WSU crossbred and commercial cows. Cholesterol content of longissimus dorsi muscle was not influenced by sire, cow herd or sex (P>0.05).     

Modifications to methods for the enumeration and detection of injured Escherichia coli O157:H7 in foods

Reliable methods are required for the detection and enumeration of potentially injured E. coli O157 in food in the presence of outnumbering competing bacteria. Selective agents can prevent or inhibit the recovery and subsequent multiplication of injured cells and direct inoculation, either into selective enrichment broths or onto selective agar plates is still used in many methods for E. coli O157 detection and enumeration. When compared with tryptone soya agar (TSA), sorbitol MacConkey agar (SMAC) was shown to underestimate the concentration of viable E. coli O157:H7 subjected to low pH and high NaCl concentration. Using a resuscitation stage on TSA followed by membrane transfer to SMAC improved recovery to levels obtained on TSA. The membrane method was used to monitor the numbers of artificially contaminated E. coli O157:H7 during the fermentation of a meat product and demonstrated better survival when compared to counts on SMAC. Six rapid methods for the detection of E. coli O157 in food (BAX E. coli O157, Reveal 8 E. coli O157-H7 screening test, VIP EHEC, VIDAS E. coli O157 (ECO), EHEC-Tek and Tecra E. coli O157 visual immunoassay), were evaluated using beetburgers, parsley and fermented meat artificially contaminated with injured cells. Methods using direct selective enrichment, with or without an elevated incubation temperature gave false-negative results. The incorporation of a non-selective pre-enrichment medium improved the detection rates of these assays by up to ten fold.     

Subsecond adsorption and desorption of dopamine at carbon-fiber microelectrodes

High-repetition fast-scan cyclic voltammetry and chronoamperometry were used to quantify and characterize the kinetics of dopamine and dopamine-o-quinone adsorption and desorption at carbon-fiber microelectrodes. A flow injection analysis system was used for the precise introduction and removal of a bolus of electroactive substance on a sub-second time scale to the disk-shaped surface of a microelectrode that was fabricated from a single carbon fiber (Thornel type T650 or P55). Pretreatment of the electrode surfaces consisted of soaking them in purified isopropyl alcohol for a minimum of 10 min, which resulted in S/N increasing by 200-400% for dopamine above that for those that were soaked in reagent grade solvent. Because of adsorption, high scan rates (2,000 V/s) are shown to exhibit equivalent S/N ratios as compared to slower, more traditional scan rates. In addition, the steady-state response to a concentration bolus is shown to occur more rapidly when cyclic voltammetric scans are repeated at short intervals (4 ms). The new methodologies allow for more accurate determinations of the kinetics of neurotransmitter release events (10-500 ms) in biological systems. Brain slice and in vivo experiments using T650 cylinder microelectrodes show that voltammetrically measured uptake kinetics in the caudate are faster using 2,000 V/s and 240 Hz measurements, as compared to 300 V/s and 10 Hz.     

Changes in rainfall chemistry and airborne particulates during a period of major local industrial change

Data from a site in central Scotland were used to quantify the changes in rainfall quality from 1989 to 1998. During this period there have been major changes in industrial activities in the area, particularly the decline in local steel-making and steel-processing activities. Many element concentrations in rainfall decreased over time in parallel with the phased reduction in the activity of local pollutant sources. Trend analyses of the rainfall data identified that the most significant responses have been the lower concentrations of Ca, SO4-S and Mn. There was also a dramatic decline in the capture of airborne particulates by the interception rainfall gauges. Particulates were found to contain mainly hematite, magnetite and quartz, that is similar to what would be expected to be derived from the neighbouring steel industries. The eventual disappearance of these particulates and the responses in rainfall quality match the timescale for the decline and closure of some of the potential sources of pollutants.     

Dynamical mechanism for sharp orientation tuning in an integrate-and-fire model of a cortical hypercolumn

Orientation tuning in a ring of pulse-coupled integrate-and-fire (IF) neurons is analyzed in terms of spontaneous pattern formation. It is shown how the ring bifurcates from a synchronous state to a non-phase-locked state whose spike trains are characterized by clustered but irregular fluctuations of the interspike intervals (ISIs). The separation of these clusters in phase space results in a localized peak of activity as measured by the time-averaged firing rate of the neurons. This generates a sharp orientation tuning curve that can lock to a slowly rotating, weakly tuned external stimulus. Under certain conditions, the peak can slowly rotate even to a fixed external stimulus. The ring also exhibits hysteresis due to the subcritical nature of the bifurcation to sharp orientation tuning. Such behavior is shown to be consistent with a corresponding analog version of the IF model in the limit of slow synaptic interactions. For fast synapses, the deterministic fluctuations of the ISIs associated with the tuning curve can support a coefficient of variation of order unity.     

Genomic organization of a voltage-gated Na+ channel in a hydrozoan jellyfish: insights into the evolution of voltage-gated Na+ channel genes

Voltage-gated Na+ channels are responsible for fast propagating action potentials. The structurally simplest animals known to contain rapid, transient, voltage-gated currents carried exclusively by Na+ ions are the Cnidaria. The Cnidaria are thought to be close to the origin of the metazoan radiation and thus are pivotal organisms for studying the evolution of the Na+ channel gene. Here we describe the genomic organization of the Na+ channel alpha subunit, PpSCN1, from the hydrozoan jellyfish, Polyorchis penicillatus. We show that most of the 20 intron sites in this diploblast are conserved in mammalian Na+ channel genes, with some even shared by Ca2+ channels. One of these conserved introns is spliced by a rare U 12-type spliceosome. Such conservation places the origin of the primary exon arrangement of Na+ channels and different intron splicing mechanisms to at least the common ancestors of diploblasts and triploblasts, approximately 600 million-1 billion years ago.