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M Hausmann

Publications and source records attributed to M Hausmann.

At least 55 records · Page 3Linked to original sources
PubMed1997

Three genes of a motility operon and their role in flagellar rotary speed variation in Rhizobium meliloti.

The peritrichous flagella of Rhizobium meliloti rotate only clockwise and control directional changes of swimming cells by modulating flagellar rotary speed. Using Tn5 insertions, we have identified and sequenced a motility (mot) operon containing three genes, motB, motC, and motD, that are translationally coupled. The motB gene (and an unlinked motA) has been assigned by similarity to the Escherichia coli and Bacillus subtilis homologs, whereas motC and motD are new and without known precedents in other bacteria. In-frame deletions introduced in motB, motC, or motD each result in paralysis. MotD function was fully restored by complementation with the wild-type motD gene. By contrast, deletions in motB or motC required the native combination of motB and motC in trans for restoring normal flagellar rotation, whereas complementation with motB or motC alone led to uncoordinated (jiggly) swimming. Similarly, a motB-motC gene fusion and a Tn5 insertion intervening between motB and motC resulted in jiggly swimming as a consequence of large fluctuations in flagellar rotary speed. We conclude that MotC biosynthesis requires coordinate expression of motB and motC and balanced amounts of the two gene products. The MotC polypeptide contains an N-terminal signal sequence for export, and Western blots have confirmed its location in the periplasm of the R. meliloti cell. A working model suggests that interactions between MotB and MotC at the periplasmic surface of the motor control the energy flux or the energy coupling that drives flagellar rotation.

Amino Acid Sequence↗
PubMed1997

Optimized Fast-FISH with alpha-satellite probes: acceleration by microwave activation.

It has been shown for several DNA probes that the recently introduced Fast-FISH (fluorescence in situ hybridization) technique is well suited for quantitative microscopy. For highly repetitive DNA probes the hybridization (renaturation) time and the number of subsequent washing steps were reduced considerably by omitting denaturing chemical agents (e.g., formamide). The appropriate hybridization temperature and time allow a clear discrimination between major and minor binding sites by quantitative fluorescence microscopy. The well-defined physical conditions for hybridization permit automatization of the procedure, e.g., by programmable thermal cycler. Here, we present optimized conditions for a commercially available X-specific alpha-satellite probe. Highly fluorescent major binding sites were obtained for 74 degrees C hybridization temperature and 60 min hybridization time. They were clearly discriminated from some low fluorescent minor binding sites on metaphase chromosomes as well as in interphase cell nuclei. On average, a total of 3.43 +/- 1.59 binding sites were measured in metaphase spreads, and 2.69 +/- 1.00 in interphase nuclei. Microwave activation for denaturation and hybridization was tested to accelerate the procedure. The slides with the target material and the hybridization buffer were placed in a standard microwave oven. After denaturation for 20 sec at 900 W, hybridization was performed for 4 min. at 90 W. The suitability of a microwave oven for Fast-FISH was confirmed by the application to a chromosome 1-specific alpha-satellite probe. In this case, denaturation was performed at 630 W for 60 sec and hybridization at 90 W for 5 min. In all cases, the results were analyzed quantitatively and compared to the results obtained by Fast-FISH. The major binding sites were clearly discriminated by their brightness.

Chromosomes↗
PubMed1996

Optimization of Fast-FISH for alpha-satellite DNA probes.

It has been shown for several highly repetitive DNA probes that the newly introduced Fast-FISH (fast-fluorescence in situ hybridization) technique is well suited for quantitative microscopy. The advantage of omitting denaturing chemical agents (e.g., formamide) in the hybridization buffer results in a short hybridization time and a considerable reduction of the number of subsequent washing steps. Choosing the appropriate hybridization temperature and time allows to clearly discriminate major and minor binding sites by quantitative fluorescence microscopy. To further optimize the procedure with reference to reproducibility, a fully programmable thermal-cycler was applied for thermal de- and renaturation. Here, the optimized renaturation conditions for two commercially available alpha-satellite probes (specific for chromosomes 1 and X) are described. For the Boehringer chromosome-1-specific DNA probe, two highly fluorescent binding sites were obtained for 72 degrees C hybridization temperature and 60 min hybridization time. For the Oncor chromosome-X-specific DNA probe, the optimal conditions were found at 74 degrees C and 60 min hybridization time. In both cases the major binding sites were clearly discriminated from only a few weakly fluorescent minor binding sites on metaphase spreads as well as in interphase cell nuclei.

DNA Probes↗
PubMed1996

Comparative genomic hybridization imaging by the one-chip true-color CCD camera kappa CF 15 MC.

In comparative genomic hybridization (CGH), quantitative measurement of fluorescence intensity ratios on metaphase chromosomes is the basic method for detecting copy number changes in the test DNA. The microscope images are usually acquired by high-resolution, highly sensitive, black and white (B&W) CCD cameras. This requires subsequent recording of the different color images using appropriate filter combinations for excitation and emission. We describe an alternative approach using the one-chip true-color CCD camera Kappa CF 15 MC and an Omega triple-bandpass filter for simultaneous registration of the three dyes Texas red, FTTC, and DAPI. A detailed examination of the imaging properties of the system was performed. The camera response in the three color planes R, G, and B was evaluated, and calibration factors for image correction were calculated. An error estimate is given. A complete computer program for the image analysis of CGH experiments has been developed for an 80486 PC, using the commercially available software package Optimas as the basis for image recording. Examples that confirm the suitability of the system for ratio imaging in CGH are presented. The results were compared with others obtained from the same microscope slides using an established setup consisting of a B&W CCD camera (Photometrics) and a software program based on the TCL software package and run on a Macintosh Quadra 950. The results obtained using the two different systems were found to correspond closely.

Calibration↗
PubMed1996

[Local efficiency of percutaneous radiotherapy in lung cancer. Analysis of 215 repeated bronchoscopies in relation to applied radiation dosage].

PURPOSE: During a locoregional radiotherapy with curative attempts of lung cancer patients bronchoscopic examinations with biopsies and/or cytologic lavages were repeated to assess the accuracy of limiting the total dose to 60 Gy. In order of the applied dose macroscopic changements of the endoluminal tumor and microscopic elimination should be made out. The correlation between macro- and microscopical regression should allow a statement about reliability of single results. The clinical course and a conventional thoracic X-ray examination seemed to be a to large-meshed screen to evaluate the effect at the end of therapy. The aim was to improve the criterias of success and to adapt and optimize the radiation dose individually. PATIENTS AND METHODS: The prospective, together with the pneumologists, defined treatment concept included the repetition of bronchoscopic evaluations after the application of 60 Gy and 80 Gy. These radiation doses from 60 Gy up to 80 Gy have been given with a shrinking-field technique to the mediastinum and the primary. In order to record statistically the optical tumor changements we were urged to create a so-called bT-score. The structure of this score was orientated towards the periphery of the tracheobronchial tree. RESULTS: Hundred and forty-four patients with endoscopically and histologically verified bronchogenic carcinomas were treated. On the subjects 215 re-bronchoscopies accomplished with biopsies were performed and allowed to analyze the macro- and microscopical behavior under treatment. A histological/cytological elimination of tumor was achieved after 60 Gy in 35.1%, after 80 Gy in 62.3%. Macroscopically no tumor was visible after 60 Gy in 43.6%, after 80 Gy in 82%. A correlation between identical micro- and macroscopical observations was only seen in 61%, respectively in 71%. CONCLUSIONS: The escalation of the radiation dose from 60 Gy up to 80 Gy with shrinked fields could increase the local tumor sterilization rate by 1.8 times from 35.1% to 62.3%. The refining and completion of usually known parameters by endoscopical and histological examinations seems to be an acceptable way to define individual radiation doses. The quality of the performed therapy can be better determined. A predestination of the total dose to a limit of 60 Gy does not ensure a macro- and microscopical elimination of the tumor and may be inferior to an individually adaptation of the dose.

Adenocarcinoma↗
PubMed1995

Granzyme A-deficient mice retain potent cell-mediated cytotoxicity.

Granzyme A, a granule-associated serine proteinase of activated cytotoxic T cells and natural killer cells, has been reported to play a critical role in DNA fragmentation of target cells. To address the question of the biological role of granzyme A, we have now generated a granzyme A-deficient mouse mutant by homologous recombination. Western blot analysis, enzyme assays and reverse transcription-PCR confirmed the absence of granzyme A in activated T cells. In addition, deletion of granzyme A does not alter the expression patterns of other granule components, such as granzymes B-G and perforin. Granzyme A-deficient mice are healthy and show normal hematopoietic development. Most notably, their in vitro- and ex vivo-derived cytotoxic T cells and natural killer cells are indistinguishable from those of normal mice in causing membrane disruption, apoptosis and DNA fragmentation in target cells. Furthermore, granzyme A-deficient mice readily recover from both lymphocytic choriomeningitis virus and Listeria monocytogenes infections and eradicate syngeneic tumors with kinetics similar to the wild-type strain. These results demonstrate that granzyme A does not play a primary role in cell-mediated cytotoxicity, as has been assumed previously.

Amino Acid Sequence↗
PubMed1995

Continuous focusing of biological particles by continuous immuno magnetic sorter: technique and applications.

We have developed a device for continuous deviation-mode, open-gradient fractionation of strongly magnetizable particles based on an electrophoresis counter-flow chamber. A mixture of magnetically labeled and nonlabeled particles can be injected into a given continuously flowing chamber buffer. The particles pass the inhomogeneous magnetic field of the open-gradient electromagnet in two narrow streams. According to the magnetic moments, induced by the magnetic field, magnetically labeled particles are deviated. The nonlabeled particles pass the magnetic field with negligible interaction. The deviated particles are focused into a stream that is completely separated from the streams of the nondeviated particles. The streams are fractionated by the counter-flow technique and collected in different vials. A high sorting purity, depending only on the specificity of the antibodies or similar labeling techniques, and a high through-put rate of up to 10(9) particles per hour was achieved. This was experimentally shown both by test particles and blood cells. The vital conditions of these blood cells were maintained by magnetic sorting.

B-Lymphocytes↗
PubMed1995

Familial juvenile polyposis: patterns of recurrence and implications for surgical management.

BACKGROUND: Familial juvenile polyposis predisposes to the development of carcinoma of the colon. Optimum surgical management and recommended surveillance of affected individuals are still being defined. STUDY DESIGN: A retrospective review of experience with a kindred identified in 1988 was carried out. RESULTS: Of 34 living members, 15 have been investigated, and histologically typical juvenile polyps were found in 11. In each instance, polyps were most numerous in the right colon, with few polyps in the descending colon and none in the rectum. Eight patients have had subtotal colectomies with ileorectal anastomoses; the remaining patients were managed by polypectomy (with one recurrence after ten years). In addition to juvenile polyps, polyps with adenomatous or villous elements were identified in three patients. One of these patients had invasive adenocarcinoma in a large mixed polyp of the cecum. Two patients with polyps had coexisting carcinoma of the stomach. All patients have been followed up with periodic upper and lower gastrointestinal endoscopy. Polyps have recurred in the rectal remnants of three patients at a mean of 36 months after subtotal colectomy. Two patients have undergone conversion to total proctocolectomy with ileoanal anastomosis and J pouch; one patient was found to have juvenile polyps in the pouch 40 months after surgery. CONCLUSIONS: Despite the preponderance of right-sided polyps at initial diagnosis, the rapid recurrence of polyps after subtotal colectomy argues in favor of performing proctocolectomy with preservation of anal sphincter function (restorative proctocolectomy) at the time of initial surgery. Patients with a small number of polyps may choose instead to undergo periodic colonoscopy with colonoscopic polypectomy. An algorithm for surveillance and follow-up is proposed.

Adenocarcinoma↗
PubMed1994

Rapid fluorescence in situ hybridization with repetitive DNA probes: quantification by digital image analysis.

Fluorescence in situ hybridization (FISH) has become an important tool not only in cytogenetic research but also in routine clinical chromosome diagnostics. Here, results of a quantification of fluorescence signals after in situ hybridization with repetitive DNA probes are reported using a non-enzymatic hybridization technique working with a buffer system not containing any formamide or equivalent chemical denaturing agents. Following simultaneous denaturation of both cells and DNA probes, the renaturation time was reduced to less than 30 min. For one of the DNA probes reasonable FISH-signals were even achieved after about 30 s renaturation time. In addition, the number of washing steps was reduced drastically. As a model system, two repetitive DNA probes (pUC 1.77, D15Z1) were hybridized to human metaphase spreads and interphase nuclei obtained from peripheral blood lymphocytes. The probes were labelled with digoxigenin and detected by FITC-anti-digoxigenin. The hybridization time was reduced step by step and the resulting fluorescence signals were examined systematically. For comparison the pUC 1.77 probe was also hybridized according to a FISH protocol containing 50% formamide. By renaturation for 2 h and overnight two FISH signals per nucleus were obtained. Using shorter renaturation times, no detectable FISH signals were observed. Quantification of the FISH signals was performed using a fluorescence microscope equipped with a cooled colour charge coupled device (CCD) camera. Image analysis was made interactively using a commercially available software package running on a PC (80486). For the pUC 1.77 probe the major binding sites (presumptive chromosomes 1) were clearly distinguished from the minor binding sites by means of the integrated fluorescence intensity. For the two (pUC 1.77) or four (D15Z1) brightest spots on the metaphase spreads and in the interphase nuclei hybridized without formamide, integrated fluorescence intensity distributions were measured for different renaturation times (0.5, 15, 30 min). The intra-nuclear variation in the intensity of the two brightest in situ hybridization spots appeared to be slightly higher (CV between 16 and 32%) than the corresponding variation in the metaphase spreads (CV between 10 and 19%). For the D15Z1 probe FISH signals were detected after hybridization without formamide and 15 min and 30 min renaturation. Always four bright spots were visible and tentatively assigned on the metaphase spreads (presumptive chromosome 15 and 9). The intensity variation of each pair of homologues in a metaphase spread showed a CV of 14 or 15%, respectively, for the presumptive chromosome 15, and 8 or 9%, respectively, for the presumptive chromosome 9.

Analog-Digital Conversion↗
PubMed1994

[Thoracoscopic pleurodesis in spontaneous pneumothorax].

The immediate effects and long-term results are reported of thoracoscopic pleurodesis in 225 patients (158 men, 67 women) treated for persistent or recurrent spontaneous pneumothorax. The procedure was performed by combined local and neurolept analgesia with direct visual exploration of the pleural space through a rigid thoracoscope. The technique included electrocoagulation of small pleural blebs, followed by regional application of fibrin and insufflation of talc powder. The main indications were a first event which persisted more than 7 days despite chest-tube suction drainage in 27% (n = 61) or a recurrent event in 73% (n = 164). The procedure provided primary success in 96.4% of the patients. Only 8 patients (3.6%) required surgical intervention including parietal pleurectomy. Perioperative complications were pharmacologically induced respiratory failure (n = 5), generalized subcutaneous emphysema (n = 8), bleeding by cutting adhesions (n = 5) and Horner's syndrome (n = 2). However, no fatal complications occurred which could be ascribed to the procedure and all patients were discharged from the hospital after an average of 12.3 days except one who died of pulmonary embolism 5 days after thoracoscopy. Long-term follow-up over a mean period of 4.1 years revealed an ipsilateral recurrence rate of 10.2% (n = 24), 16% of the patients complained of sporadic pains at the site of insertion, 51% still had diffuse thoracic pains and 2.4% reported occasional attacks of dyspnea. Spirometric lung function tests showed normal values in 89%. The immediate and longterm results show thoracoscopic pleurodesis with fibrin and talcum to be a safe and effective method for treatment of patients with persistent or recurrent pneumothorax.

Adolescent↗
PubMed1994

A rapid FISH technique for quantitative microscopy.

Results of quantitative microscopy for fluorescence in situ hybridization (FISH) signals with repetitive DNA probes (pUC 1.77 and D15Z1) are reported. A nonenzymatic hybridization technique was applied using fluorescein-12-dUTP labeled DNA probes and a buffer system not containing any formamide or equivalent chemical denaturing agents. Following thermal denaturation, the renaturation time was reduced to less than 30 min. The number of wash steps was reduced to one. For the pUC 1.77 probe, the major binding sites (chromosome 1) were distinguished from the minor binding sites by means of fluorescence intensity and spot size. The intensity variation of the two brightest FISH spots (major binding sites) in the same metaphase was 19% for 15 min renaturation time and 16% for 30 min renaturation time. For the D15Z1 probe, generally four bright spots were visible and tentatively assigned according to chromosome length and centromere position (chromosomes 15 and 9). The intensity variation of each two homologues in the same metaphase spread showed a coefficient of variation of 47% (15 min) and 22% (30 min) for chromosome 15, and 19% (15 min) and 15% (30 min) for chromosome 9. The results indicate that the applied technique can considerably accelerate the FISH procedure and is suited for quantitative microscopy.

DNA Probes↗
PubMed1994

[Yellow nail syndrome].

The yellow nail syndrome, first described in 1964 by Samman and White, consists of yellowish, dystrophic nails, lymphedema and pleural effusion. We present the clinical findings of a 53 years old woman who suffered of this very rare syndrome. The literature review reveals the different clinical features which were reported to be associated with the yellow nail syndrome. The pathogenesis of this autosomal-dominantly inherited syndrome consists of a dysfunction of the lymphatic drainage system, but details remain unknown. Up to now a causative treatment does not exist but topic tocopherol may improve the aspect of involved nails. A new study reports of good effects of oral zinc-application to nails and lymphedema.

Bronchiectasis↗
PubMed1993

Time-optimized analysis of slit-scan chromosome profiles on a general-purpose personal computer.

Slit-scan flow cytometry provides a method to analyze large numbers of metaphase chromosomes in a relatively short time due to morphological features. The high detection rate requires fast computing for on-line analysis. Up to now, this has been achieved using special-purpose computers, parallel systems or other complex hardware. Here, we describe an algorithm that can be implemented on a general-purpose personal computer. Digitized chromosome profiles can be classified by several criteria especially for the detection of chromosome abnormalities in biological dosimetry. A data set of approximately 4600 profiles was used. Programming in assembler results in an average computing time of about 600 microseconds per profile. Thus on-line evaluation of slit-scanning data appears to become feasible for many flow cytometers running nowadays.

Algorithms↗
PubMed1993

[Thoracoscopic lung biopsy in diagnosis of interstitial lung diseases].

Thoracoscopic lung biopsy is accepted to be an appropriate method in diagnosing interstitial lung diseases when lung tissue examination is needed for accurate diagnosis. In our study of 30 patients with chronic interstitial lung diseases a high sensitivity of 97% was reached similar to the yield reported from open lung biopsy techniques. The manyfold described handicap of long lasting pleural fistula after thoracoscopic biopsies could be prevented in most cases by sealing the biopsy site with a fibrin layer. Thus the mean pleural drainage-time was reduced to 3.7 +/- 1.0 days and only 3 out of the 30 patients required a prolonged drainage up to 11 days. There were only a few minor complications except in one patient who developed an etiologically unclear pulmonary edema shortly after the procedure. However the results obtained by this modified bioptic technique are encouraging and confirm again the efficacy and safety of thoracoscopic lung biopsy in diagnosing interstitial lung diseases.

Biopsy↗
PubMed1992

A tilting device for three-dimensional microscopy: application to in situ imaging of interphase cell nuclei.

The resolution of an optical microscope is considerably less in the direction of the optical axis (z) than in the focal plane (x-y plane). This is true of conventional as well as confocal microscopes. For quantitative microscopy, for instance studies of the three-dimensional (3-D) organization of chromosomes in human interphase cell nuclei, the 3-D image must be reconstructed by a point spread function or an optical transfer function with careful consideration of the properties of the imaging system. To alleviate the reconstruction problem, a tilting device was developed so that several data sets of the same cell nucleus under different views could be registered. The 3-D information was obtained from a series of optical sections with a Zeiss transmission light microscope Axiomat using a stage with a computer-controlled stepping motor for movement in the z-axis. The tilting device on the Axiomat stage could turn a cell nucleus through any desired angle and also provide movement in the x-y direction. The technique was applied to 3-D imaging of human lymphocyte cell nuclei, which were labelled by in situ hybridization with the DNA probe pUC 1.77 (mainly specific for chromosome 1). For each nucleus, 3-D data sets were registered at viewing angles of 0 degrees, 90 degrees and 180 degrees; the volumes and positions of the labelled regions (spots) were calculated. The results also confirm that, in principle, any angle of a 2 pi geometry can be fixed for data acquisition with a high reproducibility. This indicates the feasibility of axiotomographical microscopy of cell nuclei.

Cell Nucleus↗
PubMed1992

An adapter for defined sample volumes makes it possible to count absolute particle numbers in flow cytometry.

A device is described which makes it possible to count absolute particle (cell) numbers per volume by flow cytometry. It can easily by adapted to several types of flow cytometers, especially to the Coulter EPICS V and EPICS 750 series. A volume adapter has been installed in place of the normal sample handling system without any further modifications of the instrument or the data acquisition program. The adapter consists of a special pipette with two opto-electronic detectors for the beginning and end of the measuring period. These switch on/off a shutter for the illuminating laser beam so that acquisition of the data is controlled indirectly. Sample volumes of 50 microliters were measured at flow rates up to 10(3) particles/s. Calibration beads as well as blood cells were enumerated according to FALS (forward angle light scatter), to SSC (90 degrees light scatter), and to fluorescence parameters. The results were compared to the evaluation made on a Coulter counter or in a Neubauer chamber of a light microscope. Using a concentration of 1 x 10(5)-5 x 10(5) particles/ml, the absolute numbers of particles were determined with a high reproducibility and an estimated error rate of 2-5%.

Antibodies, Monoclonal↗
PubMed1991

Background and peak evaluation of one parameter flow karyotypes on a PC/AT computer.

Flow cytometry has become a fast, quantitative method for the classification of metaphase chromosomes in suspension (flow karyotyping) stained with fluorescent dyes. Such a flow karyotype (frequency distribution of the fluorescence signals) consists of several peaks. The peak pattern characterizes the analyzed chromosome complement. In many cases flow karyotypes contain a continuum of an unspecific background deriving from chromosome fragments or chromosome aggregates. For the quantitative evaluation of a flow karyotype this background has to be subtracted by a suitable background function. In this approach the application of chi 2-functions is described. The feasibility of this method to flow karyotypes has been concluded from a computer simulation of chromosome breaking under different conditions. In spite of the rather rough assumptions of the model compared to the complex reasons that influence chromosome breaking, the chi 2-function fits the background better than the exponential function in current use. The approximation of a Gaussian distribution function by the chi 2-function also makes it possible to use the same subtraction procedure for chromosome aggregates. The procedure was tested for isolated chromosomes of Chinese hamster cell lines under different states of breaking. For further evaluation of one parameter flow karyotypes a setup of computer routines has been developed for PC/AT and compatible computer systems. Different peak values of these flow karyotypes can be determined (e.g. peak mean, standard deviation, absolute and relative peak area etc.). The applied method is to fit Gaussian curves to each peak of an experimentally measured histogram by using an interactive program. Fluctuations depending on 'noise' may be suppressed by a 'k-nearest-neighbours' smoothing procedure.

Animals↗