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J Hampl

Publications and source records attributed to J Hampl.

At least 19 recordsLinked to original sources
PubMed2004

Some basic parameters of microspheres fabricated from a branched oligoester by a rapid procedure.

Microspheres were prepared from a branched copolymer of DL-lactic acid with mannitol containing native albumin and albumin labelled with fluorescein isothiocyanate, using a rapid method of distribution of methylformate as the solvent of the copolymer from the intermediate phase of the multiple w/o/w emulsion. The primary w/o emulsion was prepared by the method of homogenization with a turbine or, alternatively, by the method of dispersion with ultrasound in modified vessels. Different additives in the external aqueous phase, such as polyvinyl alcohol or the gelatin hydrolyzate as emulsifiers were tested. Ammonium sulphate, methylformate or ethyl acetate were used as moderators of solidification of microspheres. The effect of these selected formulation parameters on the size, encapsulation efficiency, yield of microspheres and on the course of the BSA and FITC-BSA release in vitro conditions were examined.

Acetates↗
PubMed2001

Upconverting phosphor reporters in immunochromatographic assays.

Immunochromatographic assays have become popular diagnostic tools in a variety of settings because they are sensitive, fast, and easy to use. Here, we describe the use of a novel reporter, upconverting phosphors (UCP), in this assay format. UCP are submicron-sized, inorganic crystals that are excited with infrared light and that emit photons in the visible range depending on the ion composition of the crystal. Using human chorionic gonadotropin (hCG) as a model analyte to describe the properties of phosphors in immunochromatographic assays, a detection limit of 10 pg hCG in a 100-microl sample has been achieved on a regular basis, with occasional detection of 1 pg hCG. This represents at least a 10-fold improvement over conventional reporter systems such as colloidal gold or colored latex beads. Quantitation of analytes is possible over at least 3 orders of magnitude. Furthermore, an example is given of how UCP can be used for analyte multiplexing using a two-plexed wick for the detection of mouse IgG and ovalbumin. Thus, UCP lateral flow assays can be used for applications that are currently limited by assay sensitivity, and they can increase the probability of a diagnosis by verifying the presence of several analytes in the same sample.

Chorionic Gonadotropin↗
PubMed2000

A population of murine gammadelta T cells that recognize an inducible MHC class Ib molecule.

Although gammadelta T cells are implicated in regulating immune responses, gammadelta T cell-ligand pairs that could mediate such regulatory functions have not been identified. Here, the expression of the major histocompatibility complex (MHC) class Ib T22 and the closely related T10 molecules is shown to be activation-induced, and they confer specificity to about 0.4% of the gammadelta T cells in normal mice. Thus, the increased expression of T22 and/or T10 might trigger immunoregulatory gammadelta T cells during immune responses. Furthermore, the fast on-rates and slow off-rates that characterize this receptor/ligand interaction would compensate for the low ligand stability and suggest a high threshold for gammadelta T cell activation.

Animals↗
PubMed1999

The specificity of a weak gamma delta TCR interaction can be modulated by the glycosylation of the ligand.

The gamma delta T cell clone LBK5 recognizes the MHC molecule IEk. Here, we demonstrate that the affinity of this interaction is weaker than those typically reported for alpha beta TCRs that recognize peptide/MHC complexes. Consistent with our previous finding that peptide bound to the IE molecule does not confer specificity, we show that the entire epitope for LBK5 is contained within the polypeptide chains of the molecule, centered around the polymorphic residues 67 and 70 of the IE beta-chain. However, LBK5 recognition is profoundly influenced by the N-linked glycosylation at residue 82 of the IE alpha-chain. Since infected, stressed, or transformed cells often change the posttranslational modifications of their surface glycoproteins, this finding suggests a new way in which gamma delta T cell Ag recognition can be regulated.

Animals↗
PubMed1998

Evidence that TSG101 aberrant transcripts are PCR artifacts.

Critical analysis of the data published so far concerning the TSG101 gene revealed some inconsistencies leading us to its re-evaluation in 80 breast, brain, colon, and neuroectodermal tumors and 37 normal tissue specimens. In this study, the occurrence of TSG101 cDNA aberrant transcripts was verified, but in addition we made observations that are in apparent conflict with the aberrant splicing theory supposed as the underlying mechanism for transcript formation: the location of most deletion breakpoints within exons and nonconformity of these putative splice sites with the highly conserved GT-AG rule, detection of insertions as well as nonreproducible and highly variable results in repeated RT-nested PCRs. Furthermore, we found that reamplification of full-length TSG101 cDNA products leads to the generation of deleted transcripts. In summary, for the first time we provide evidence that the acquired TSG101 transcripts are caused by PCR artifacts.

Artifacts↗
PubMed1998

Initiation of signal transduction through the T cell receptor requires the multivalent engagement of peptide/MHC ligands [corrected].

While much is known about intracellular signaling events in T cells when T cell receptors (TCRs) are engaged, the mechanism by which signaling is initiated is unclear. We have constructed defined oligomers of soluble antigen-major histocompatibility complex (MHC) molecules, the natural ligands for the TCR. Using these to stimulate specific T cells in vitro, we find that agonist peptide/MHC ligands are nonstimulatory as monomers and minimally stimulatory as dimers. Similarly, a partial-agonist ligand is very weakly active as a tetramer. In contrast, trimeric or tetrameric agonist ligands that engage multiple TCRs for a sustained duration are potent stimuli. Ligand-driven formation of TCR clusters seems required for effective activation and helps to explain the specificity and sensitivity of T cells.

Amino Acid Sequence↗
PubMed1998

Ligand recognition by alpha beta T cell receptors.

While still incomplete, the first data concerning the biochemistry of T cell receptor-ligand interactions in cell-free systems seem to have considerable predictive value regarding whether a T cell response is strong or weak or suppressive. This data will help considerably in elucidating the mechanisms behind T cell responsiveness. Also of great interest are the first structures of T cell receptor molecules and, particularly, TCR-ligand complexes. These appear to confirm earlier suggestions of a fixed orientation for TCR engagement with peptide/MHC and should form the basis for understanding higher oligomers, evidence for which has also just emerged. We conclude with an analysis of the highly diverse CDR3 loops found in all antigen receptor molecules and suggest that such regions form the core of both TCR and antibody specificity.

Animals↗
PubMed1997

The putative tumor suppressor gene FHIT at 3p14.2 is rarely affected by loss of heterozygosity in primary human brain tumors.

To elucidate the role of the recently identified FHIT gene, located at 3p14.2 in human brain tumor carcinogenesis, a total of 259 tumors were analyzed for loss of heterozygosity (LOH) at microsatellite loci D3S1313, D3S1234, D3S1300, and D3S1481. In primary brain tumors, LOH was detected at a frequency of 8.4% (n = 214). Low-grade gliomas exhibited insignificantly lower LOH rates in comparison to high-grade gliomas (5.3%, n = 19, versus 11.1%, n = 90). Notably, no allelic loss was observed in 12 recurrent glioblastomas analyzed in comparison to their corresponding primary tumor lesions and in two astrocytomas with progression to higher grades of malignancy. Our data indicate that allelic loss of the FHIT gene is neither a critical event in carcinogenesis of primary brain tumors nor tumor grade-associated in astrocytic tumors. In contrast, observed LOH rate for brain metastases was as high as 54.5% (n = 45), in accordance with data thus far accumulated from analyses of corresponding primary tumors.

Acid Anhydride Hydrolases↗
PubMed1997

T cell receptor biochemistry, repertoire selection and general features of TCR and Ig structure.

T cell recognition is a central event in the development of most immune responses, whether appropriate or inappropriate (i.e. autoimmune). We are interested in reducing T cell recognition to its most elemental components and relating this to biological outcome. In a model system involving a cytochrome c-specific I-Ek restricted T cell receptor (TCR) derived from the 2B4 hybridoma, we have studied the interaction of soluble TCR and soluble peptide-MHC complexes using surface plasmon resonance. We find a striking continuum in which biological activity correlates best with the dissociation rate of the TCR from the peptide-MHC complex. In particular, we have found that weak agonists have significantly faster off-rates than strong agonists and that antagonists have even faster off-rates. This suggests that the stability of TCR binding to a given ligand is critically important with respect to whether the T cell is stimulated, inhibited or remains indifferent. It also suggests that the phenomenon of peptide antagonists might be explained purely by kinetic models and that conformation, either inter- or intramolecular, may not be a factor. We have also studied TCR repertoire selection during the establishment of a cytochrome c response, initially using an anti-TCR antibody strategy, but more recently using peptide-MHC tetramers as antigen-specific staining reagents. These tetramers work well with either class I or class II MHC-specific TCRs and have many possible applications. Lastly, we have also tried to correlate the structural and genetic features of TCRs with their function. Recent data on TCR structure as well as previous findings with antibodies suggest that both molecules are highly dependent on CDR3 length and sequence variation to form specific contacts with antigens. This suggests a general "logic' behind TCR and Ig genetics as it relates to structure and function that helps to explain certain anomalous findings and makes a number of clear predictions.

Animals↗
PubMed1997

CD4 augments the response of a T cell to agonist but not to antagonist ligands.

The recognition of peptide variants by the T cell receptor (TCR) has revealed a wide range of possible responses. Here, using a series of CD4+ and CD4- variants of the same T cell hybridoma, we find that while the expression of CD4 converts weak agonists into full agonists, none of the antagonist peptides are efficiently recognized as agonists. Furthermore, in antagonist assays, little difference can be seen in the response of CD4+ and CD4- T cells. Together with previous work showing a marked difference in stability between TCR binding to agonist versus antagonist ligands, these data suggest that CD4 engagement occurs after a TCR-peptide/MHC complex has formed and that it requires a certain minimal half-life of the ternary complex to be fully engaged in signaling.

Amino Acid Sequence↗
PubMed1996

A TCR binds to antagonist ligands with lower affinities and faster dissociation rates than to agonists.

T lymphocyte activation is mediated by the interaction of specific TCR with antigenic peptides bound to MHC molecules. Single amino acid substitutions are often capable of changing the effect of a peptide from stimulatory to antagonistic. Using surface plasmon resonance, we have analyzed the interaction between a complex consisting of variants of the MCC peptide bound to a mouse class II MHC (Ek) and a specific TCR. Using both an improved direct binding method as well as a novel inhibition assay, we show that the affinities of three different antagonist peptide-Ek complexes are approximately 10-50 times lower than that of the wildtype MCC-Ek complex for the TCR, largely due to an increased off-rate. These results suggest that the biological effects of peptide antagonists and partial agonists may be largely based on kinetic parameters.

Amino Acid Sequence↗
PubMed1996

Enhancement of antibody response to bovine herpesvirus 1 with non-specific immunostimulants.

Stimulatory effects of aluminium hydroxide, lipopolysaccharide (LPS), muramyldipeptide (MDP), and empty liposomes on the antigenicity of inactivated bovine herpesvirus 1 were tested in mice. Compared with the standard effect of aluminium hydroxide, stronger antibody responses were observed in mice treated with empty liposomes or LPS alone, or a combination thereof. The strongest antibody response was recorded in mice treated with a combination of inactivated BHV-1, MDP and empty liposomes.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1996

Immunogenicity of infectious bovine rhinotracheitis virus (BHV-1) proteins integrated into ISCOMs or liposomes.

Dynamics and persistence of antibody responses to immunization and reimmunization of rabbits with BHV-1 ISCOMs or BHV-1 liposomes were investigated for 20 weeks and compared with those induced by a commercial vaccine against infectious bovine rhinotracheitis (IBR) or free BHV-1 protein subunits. Comparable levels of antibodies to BHV-1, determined by ELISA, were found throughout the observation period in rabbits treated with BHV-1 ISCOMs or the commercial vaccine. The immunogenicity of BHV-1 liposomes was higher than that of the free virus proteins, but lower than those of BHV-1 ISCOMs or the commercial vaccine. Virus neutralization test confirmed the best immunostimulatory effect of BHV-1 ISCOMs as far as the levels and the persistence of antibodies were concerned. The reactivities of the specific virus protein subunits with anti-BHV-1 antibodies elicited in rabbits immunized with BHV-1 ISCOMs or the commercial vaccine were compared by Western blotting. The patterns of the bands were similar, but some differences were found in their numbers and intensities.

Animals↗
PubMed1995

In vitro and in vivo efficacy of a rifampin-loaded silicone catheter for the prevention of CSF shunt infections.

Infection of cerebrospinal fluid (CSF) shunts is one of the major complications associated with their use and is usually managed by shunt removal, temporary insertion of an external drainage and implantation of a new shunt system. We have evaluated the efficacy of a rifampin-loaded silicone ventricular catheter to prevent bacterial colonization and infection in vitro and in an animal model. On the basis of an incorporation process a rifampin-loaded catheter was developed which is capable of releasing rifampin in bacteriocidal concentrations for 60 days and more. In a stationary bacterial adherence assay using S. epidermidis as test strain, the colonization resistance of the device was demonstrated. To assess the capability of the catheter to prevent CSF shunt infections, a rabbit model was developed which allowed the establishment of a reliable and reproducible CSF infection by implantation of silicone catheters into the ventricle and inoculating S. epidermidis (minimal dose 10(6) cfu) or S. aureus (minimal dose 10(3) cfu). Rifampin-loaded catheters (12 animals inoculated with S. epidermidis, 8 animals inoculated with S. aureus) were compared with non-loaded (14 animals inoculated with S. epidermidis, 19 animals inoculated with S. aureus) control catheters, and infection was documented by clinical, microbiological and histological methods. In contrast to the control group, none of the animals with rifampin-loaded catheters showed clinical signs of infection. Furthermore, in none of the materials obtained after sacrifice of the animals (catheter, brain tissue, CSF, blood) could the infecting bacteria be cultured, whereas in materials from animals with the unloaded catheter the infecting strains could always be cultured from the catheter and from surrounding brain tissue. The histological examination of catheter-adjacent tissue supported these findings. We conclude that a rifampin-loaded silicone ventricular catheter is capable of completely preventing bacterial colonization and infection by staphylococci as the main causative organisms in CSF shunt infections and should be further evaluated in clinical trials.

Animals↗
PubMed1995

Digital optical archiving of medical records in hospital information systems: results of a pilot (feasibility) study.

The paper-based medical record is a "principal repository for information" [1] and supports health care professionals in patient care, quality assessment, and often medical research. University hospitals often require expensive space for a long-time storage of medical records. The costs for operating conventional archives are high and will increase in future. In spite of these efforts, conventional archiving does not satisfy the medical needs to make available medical records for health care professionals in a systematic and timely manner.

Hospital Information Systems↗
PubMed1994

Pharmacokinetics of sodium humate in chickens.

Free or liposome-encapsulated sodium humate was administered to chickens intracardially, orally or subcutaneously and the following principal pharmacokinetic parameters were determined using the MW Pharm software for two-compartment models: elimination half-life (t1/2 el), steady state distribution volume (Vdss), blood clearance (Cl), maximal drug concentration (Cmax), time required for appearance of Cmax (tmax), area under the curve (AUC) and bioavailability (F). Blood clearance of liposome-encapsulated sodium humate was higher than that of free sodium humate regardless of the way of administration. On the other hand, the elimination half-life was longer after the extravascular than after the intracardial administration. Cmax values indicate that the penetration of sodium humate from the injection site into blood circulation is very slow. Biological availability of sodium humate also depended on the way of administration and dosage form. Aside from the intracardial administration, the highest bioavailability was found after subcutaneous administration of free sodium humate.

Animals↗