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Biomedical subjects

K E Hoffman

Publications and source records attributed to K E Hoffman.

13 recordsLinked to original sources
PubMed2000

Simultaneous detection of six biohazardous agents using a planar waveguide array biosensor.

Recently, we demonstrated that an array biosensor could be used with cocktails of fluorescent antibodies to perform three assays simultaneously on a single substrate, and that multiple samples could be analyzed in parallel. We extend this technology to demonstrate the simultaneous analysis of six samples for six different hazardous analytes, including both bacteria and protein toxins. The level of antibody cross-reactivity is explored, revealing a possible common epitope in two of the toxins. A panel of environmental interferents was added to the samples; these interferents neither prevented the detection of the analytes nor caused false-positive responses.

Bacteria↗
PubMed2000

Array biosensor for detection of biohazards.

A fluorescence-based biosensor has been developed for simultaneous analysis of multiple samples for multiple biohazardous agents. A patterned array of antibodies immobilized on the surface of a planar waveguide is used to capture antigen present in samples; bound analyte is then quantified by means of fluorescent tracer antibodies. Upon excitation of the fluorophore by a small diode laser, a CCD camera detects the pattern of fluorescent antibody:antigen complexes on the waveguide surface. Image analysis software correlates the position of fluorescent signals with the identity of the analyte. This array biosensor has been used to detect toxins, toxoids, and killed or non-pathogenic (vaccine) strains of pathogenic bacteria. Limits of detection in the mid-ng/ml range (toxins and toxoids) and in the 10(3)-10(6) cfu/ml range (bacterial analytes) were achieved with a facile 14-min off-line assay. In addition, a fluidics and imaging system has been developed which allows automated detection of staphylococcal enterotoxin B (SEB) in the low ng/ml range.

Biosensing Techniques↗
PubMed1996

Characterization of DNA synthesis during the 2-cell stage and the production of tetraploid chimeric pig embryos.

The DNA content of nuclei during the 2-cell stage as well as in presumptive tetraploid embryos was investigated. In vivo produced pig zygotes were cultured to the 2-cell stage and either monitored for cleavage to the 4-cell stage or mounted at various times post-cleavage and DNA content determined. The length of the 2-cell stage was 14.8 +/- 3.0 hr. There was a significant increase in the length of the 2-cell stage due to the time in vitro as a zygote (P < 0.001: R2 = 0.866). The DNA content increased (P < 0.05) each 2 hr postcleavage until 10 hr postcleavage. This suggested that there is a short G1 and G2 phase and a relatively long phase of DNA synthesis. Next, 2-cell stage embryos were pulsed with electricity to induce cell-to-cell fusion. Whereas only about half fused within 30 min (55%), most (96%) developed to the blastocyst stage. The DNA content of the nuclei of the embryos was consistent with them being tetraploid. A final experiment was designed to evaluate the ability of the tetraploid embryo to form a chimera with isolated inner cell mass (ICM) cells. Inner cell masses were isolated from d 6 embryos, cut into thirds, labeled with DiO (a membrane die) and injected into the perivitelline space of 4-cell-stage tetraploid embryos. Twelve of 17 formed blastocysts. In most (8/12), the ICM of the resulting blastocyst was labeled, whereas in one the only fluorescence was in the trophectoderm, and in two fluorescence was evenly distributed between the ICM and trophectoderm. These results suggest that it may be possible to create a fetus derived from ICM cells, or potentially stem cells, that has a tetraploid trophoblast.

Animals↗
PubMed1996

Evidence for the involvement of the caudal region of the periaqueductal gray in a subset of morphine-induced alterations of immune status.

This study was directed at determining whether morphine's immunomodulatory effects are mediated through the periaqueductal gray (PAG). The initial study showed that microinjection of morphine (0.0, 0.4, 4.0, or 40.0 micrograms/rat) into the lateral ventricle induces pronounced dose-dependent reductions in lymphocyte proliferation to T- and B-cell mitogens, natural killer cell cytotoxicity, and the production of interleukin-2 and interferon-gamma. In contrast, microinjection of morphine (0.0, 0.004, 0.04, 0.4, or 4.0 micrograms/rat) into the caudal aspect of the PAG induced dose-dependent alterations in natural killer cell cytotoxicity, but had no effect on lymphocyte proliferation or cytokine production. These results indicate that opioid receptors in the PAG are involved in the regulation of natural killer cell activity, but are not associated with morphine's effects on proliferation or cytokine production. A subsequent study showed that the effect of morphine in the PAG is restricted to the more caudal aspects of the PAG because microinjections of morphine into the rostral aspects do not result in any alteration of immune status. To determine that the activation of opioid receptors in the PAG is not only sufficient, but is required for morphine's effects on natural killer cell activity, N-methylnaltrexone was administered into the PAG (0, 0.0001, 0.001, or 0.01 micrograms/rat) before the systemic administration of morphine (15 mg/kg), a dose that induces pronounced alterations of natural killer cell activity. The results showed that the administration of N-methylnaltrexone directly into the PAG antagonized morphine's effects on natural killer cell activity, which indicate that activation of opioid receptors within the PAG are required for morphine to alter natural killer cell activity. Collectively, this study showed that activation of opioid receptors within the more caudal aspects of the PAG are required for morphine to induce alterations in splenic natural killer cell activity. The results also suggest that other brain regions are responsible for morphine's effect on lymphocyte proliferation and cytokine production.

Adjuvants, Immunologic↗
PubMed1994

The effects of masking on vibrotactile temporal summation in the detection of sinusoidal and noise signals.

Thresholds for the detection of vibrotactile signals of varied duration applied to the thenar eminence were measured in the absence of and in the presence of a masking stimulus. Signals were 250- and 500-Hz sinusoids and noise bursts with bandwidth limited to 250-1000 Hz. The masking stimulus was either a 250-Hz sinusoid, which was presented in phase with the signal when it was sinusoidal, or noise. Changes in threshold as a function of changes in signal duration were found which were predicted accurately from Zwislocki's theory [J. Acoust. Soc. Am. 32, 1046-1060 (1960)] of temporal summation when the signal was detected by the Pacinian channel, but not when it was detected by a non-Pacinian channel (NP). However, when either the signal or the masking stimulus or both were noise, NP thresholds were affected by changes in signal duration. Only when the signal and masking stimuli were both sinusoids were NP thresholds independent of signal duration. It is concluded that signal duration effects in the NP channel are not due to temporal integration, but rather to increases in information about the signal content provided to the subject as exposure duration is increased.

Acoustic Stimulation↗
PubMed1994

The effects of aging on information-processing channels in the sense of touch: I. Absolute sensitivity.

Thresholds for detecting vibrotactile signals of variable frequency applied to the thenar eminence of the hand by small and large contactors were measured in subjects ranging in age from 10 to 89 years. Thresholds were found to increase as a function of age, but the rate of increase was greater after than before the age of 65 years. The rate of loss of vibrotactile sensitivity was substantially greater in the P channel (mediated by Pacinian corpuscles) than in the NP I channel (mediated by rapidly adapting fibers), the NP II channel (mediated by slowly adapting type II fibers), or the NP III channel (mediated by slowly adapting type I fibers). Women were frequently found to have greater sensitivity than men.

Adolescent↗
PubMed1991

Ascorbic acid and iron metabolism: alterations in lysosomal function.

Iron is essential to cell metabolism but promotes free radical damage to membranes and lipids. Therefore, excess intracellular iron is stored within the shell of hollow ferritin molecules until needed for metabolic use. Ascorbate retards ferritin degradation and increases iron bioavailability. The vitamin stabilizes the iron cores of ferritin in cells prelabeled with 59Fe. [35S]Methionine labeling demonstrates that this enhanced stability of the iron cores results from delayed degradation of the ferritin shells. Subcellular fractionation of 59Fe-labeled cells by use of a Sepharose CL-6B column shows that ascorbate significantly delays the shift of ferritin label from the cytosolic to the lysosomal compartment. Monomeric ferritin shells in the cytoplasm gradually form clusters that bind to lysosomes. Single ferritin shells do not. Ascorbate does not affect the conversion of cytoplasmic ferritin monomers to clusters but greatly retards the autophagic uptake of ferritin clusters into lysosomes.

Animals↗
PubMed1990

The effect of scleral buckling on ocular rigidity.

In a study of enucleated human eyes, the authors investigated the effect of scleral buckling on the ocular pressure-volume relationship. Intraocular pressure was recorded continuously during intravitreal infusion of saline solution before and after the application of encircling silicone elements. Scleral buckling produced a marked reduction in ocular rigidity, with reversibility of the effect on removal of the buckling elements. Similar results were obtained during incremental intravitreal air injection. The authors propose that the greater extensibility of silicone compared with sclera and the induced alterations in ocular shape are the primary factors responsible for the observed change in ocular rigidity. The clinical implications of these findings for intravitreal gas injection are discussed.

Aged↗
PubMed1988

Guanosine triphosphate promotes the post-translational integration of opsin into the endoplasmic reticulum membrane.

Membrane integration of a nascent opsin polypeptide was examined to determine whether insertion of proteins into the endoplasmic reticulum is dependent upon energy provided by ribonucleotide triphosphate hydrolysis. A discrete-sized nascent chain was obtained by in vitro translation of a mRNA which lacked a termination codon yet encoded the first 156 residues of bovine opsin. Ribosomes bearing the newly synthesized opsin chains were post-translationally incubated with canine pancreas microsomal membrane vesicles after addition of exogenous ribonucleotides or ribonucleotide analogues. Post-translational membrane integration and glycosylation of the 156-residue nascent polypeptide was found to require either the presence of guanosine triphosphate or a nonhydrolyzable GTP analogue. ATP did not promote post-translational integration of the nascent polypeptide. Although ribonucleotide hydrolysis was not obligatorily required for integration of opsin, we observed an increase in the proportion of glycosylated opsin chains in post-translational incubations that contained hydrolyzable ribonucleotide triphosphates. We conclude that a GTP-binding protein performs an essential role during integration of opsin into the endoplasmic reticulum.

Animals↗
PubMed1986

The effects of ascorbic acid on the intracellular metabolism of iron and ferritin.

An important property of ascorbic acid is its ability to increase the availability of storage iron to chelators. To examine the mechanism of this effect, K562 cells were incubated with ascorbate, attaining an intracellular level of 1 nmol/10(7) cells. In contrast to the reductive mobilization of iron seen with isolated ferritin, ascorbate stabilized iron preincorporated into cellular ferritin. Biosynthetic labeling with [35S]methionine demonstrated that ascorbate also retarded the degradation of the ferritin protein shell. Ferritin is normally degraded in lysosomes. The lysosomal protease inhibitors leupeptin and chloroquine produced a qualitatively similar stabilization of ferritin. Ascorbate did not act as a general inhibitor of proteolysis, however, since it did not effect hemoglobin degradation in these cells. The stabilization of cellular ferritin by ascorbate was accompanied by an expansion of the pool of chelatable iron.

Animals↗