Biomedical subjects

K Aldinger

Publications and source records attributed to K Aldinger.

5 recordsLinked to original sources

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↗

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

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↗

PubMed1977

Focal nodular hyperplasia of the liver associated with high-dosage estrogens.

A patient developed benign nodular hyperplasia of the liver after long-term, high-dose estrogen therapy. A 10.5-cm benign liver nodule appeared after three years of high-dose estrogen therapy and was resected. Fourteen months later, while still receiving estrogen, a 4.5-cm nodule and multiple small nodules were seen on angiography. Four months after estrogen was discontinued, the larger liver nodule disappeared and the smaller ones decreased.

Adult↗

PubMedDate not supplied

Painting of human chromosome 8 in fifteen minutes.

The technique of chromosome-in-situ suppression (CISS)-hybridization (chromosome painting) has now been well established. However, all standard protocols so far require long renaturation times (typically 12 hours and more). Here, we describe a new, extremely fast protocol for chromosome painting using a commercially available, directly fluorescence labelled probe for chromosome 8. The hybridization conditions used omit separate preannealing procedures and denaturing chemical agents. The renaturation time required for chromosome painting was reduced to 15 minutes. In addition, most washing steps were eliminated. As a consequence, the entire painting procedure was feasible in less than half an hour.

Chromosomes, Human, Pair 8↗