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PubMed1989

Range of the solvation pressure between lipid membranes: dependence on the packing density of solvent molecules.

Well-ordered multilamellar arrays of liquid-crystalline phosphatidylcholine and equimolar phosphatidylcholine-cholesterol bilayers have been formed in the nonaqueous solvents formamide and 1,3-propanediol. The organization of these bilayers and the interactions between apposing bilayer surfaces have been investigated by X-ray diffraction analysis of liposomes compressed by applied osmotic pressures up to 6 X 10(7) dyn/cm2 (60 atm). The structure of egg phosphatidylcholine (EPC) bilayers in these solvents is quite different than in water, with the bilayer thickness being largest in water, 3 A narrower in formamide, and 6 A narrower in 1,3-propanediol. The incorporation of equimolar cholesterol increases the thickness of EPC bilayers immersed in each solvent, by over 10 A in the case of 1,3-propanediol. The osmotic pressures of various concentrations of the neutral polymer poly(vinylpyrrolidone) dissolved in formamide or 1,3-propanediol have been measured with a custom-built membrane osmometer. These measurements are used to obtain the distance dependence of the repulsive solvation pressure between apposing bilayer surfaces. For each solvent, the solvation pressure decreases exponentially with distance between bilayer surfaces. However, for both EPC and EPC-cholesterol bilayers, the decay length and magnitude of this repulsive pressure strongly depend on the solvent. The decay length for EPC bilayers in water, formamide, and 1,3-propanediol is found to be 1.7, 2.4, and 2.6 A, respectively, whereas the decay length for equimolar EPC-cholesterol bilayers in water, formamide, and 1,3-propanediol is found to be 2.1, 2.9, and 3.1 A, respectively. These data indicate that the decay length is inversely proportional to the cube root of the number of solvent molecules per unit volume.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholesterol↗
PubMed1977

Circular forms of Uukuniemi virion RNA: an electron microscopic study.

Because the ribonucleoprotein forms of the segments of the Uukuniemi virus genome have previously been characterized as circular, we examined the isolated RNAs by electron microscopy under conditions of increasing denaturation. After spreading under moderately denaturing conditions (50 or 60% formamide), 50 to 70% of the molecules were circular. Increasing the formamide concentration to 70 and 85% decreased the number of circular forms, and only linear forms were observed after incubation of the RNA at 60 degrees C for 15 min in 99% formamide. When spread from 4 M urea-80% formamide--another condition known to denature RNA--only 5 to 30% circular molecules were observed. Pretreatment of the RNA with 0.5 M glyoxal at 37 degrees C for 15 min prior to spreading from 50% formamide gave less than 5% cirucular forms. Length measurement of the molecules showed that they were not significantly degraded by any of the methods employed. The circular molecules were destroyed by treatment with pancreatic RNase, but were unaffected by DNase or proteinase K treatment. After complete denaturation of the RNA, the circles could be reformed under reannealing conditions. We conclude that the three size classes of RNA that comprise the Uukuniemi virus genome are circular molecules probably maintained in that form by base pairing between inverted complementary sequences at the 3' and 5' ends of linear molecules.

Arboviruses↗
PubMed1992

Detection of mRNA by in situ hybridisation and in northern blot analysis using oligodeoxynucleotide probes labelled with alkaline phosphatase.

AIMS: To assess whether a reduction in intensity of signal observed using an alkaline phosphatase labelled oligodeoxynucleotide probe could be explained on the basis of procedural steps rather than reduced sensitivity. METHOD: Signal intensity was assessed on in situ hybridisation for pro-opiomelanocortin (POMC) mRNA in rat pituitary and for somatostatin mRNA in human pancreas and in northern blot analysis for POMC mRNA in the presence and absence of formamide. The direct effects of formamide on the alkaline phosphatase detection step were assessed using histochemical enzyme detection in rat kidney. RESULTS: All signals were reduced in systems containing formamide. CONCLUSIONS: In the absence of formamide clear, strong signals for specific mRNAs can be obtained by in situ hybridisation and northern blot analysis using oligodeoxynucleotide probes directly labelled with alkaline phosphatase. Formamide seems to inhibit the activity of alkaline phosphatase.

Alkaline Phosphatase↗
PubMedDate not supplied

In vitro studies on the metabolic fate of mifentidine, a novel histamine H2-receptor antagonist.

The in vitro metabolism of mifentidine and several of its metabolites was studied using hepatic microsomes from seven animal species. The effects of potential enzyme inducers, inhibitors and activators were also studied. Mifentidine metabolites identified and characterised were: 4-imidazolylphenylamine (amine), 4-imidazolylphenyl-formamide (formamide), the urea derivative of mifentidine (urea) and the imidazole-hydroxylated derivative of the amine (i-OH-amine), along with three unidentified metabolites, M1, M2 and M3. Evidence for the presence of the amine, formamide, urea and i-OH-amine was obtained by comparison with authentic reference compounds: (i) HPLC retention times; (ii) UV spectra; and (iii) MS spectra of metabolites. The postulated intermediates are: carbinolimine (for formamide, amine, i-OH-amine and urea formation); formamide (for amine and i-OH-amine formation); amine (for i-OH-amine formation), and nitrone (for urea formation). One 'metabonate' of mifentidine was also identified, namely the nitro analogue of the amine. A possible prerequisite for the formation of this nitro is the corresponding hydroxylamine or nitroso compound. Cytochromes P450I and P450II were shown to be involved in the in vitro microsomal biotransformation of mifentidine, but the involvement of the flavin monooxygenase system was not proven.

Animals↗
PubMed2005

Synthesis of some derivatives of C-(1-deoxy-1-N-substituted-D-glucopyranosyl)formic acid (D-gluco-hept-2-ulopyranosonic acid) as potential inhibitors of glycogen phosphorylase.

Per-O-benzoylated derivatives (amide, methyl ester and glycinamide) of C-(1-azido-1-deoxy-alpha-D-glucopyranosyl)formic acid obtained by azide substitution in the corresponding C-(1-bromo-1-deoxy-beta-D-glucopyranosyl)formic acid derivatives were debenzoylated by the Zemplén-protocol. Per-O-benzoylated C-(1-azido-1-deoxy-alpha-D-glucopyranosyl)formamide was dehydrated by oxalyl chloride-DMF to give the corresponding nitrile, while from its reduction mixture obtained by Raney-nickel or sodium hydrogentelluride C-(1-amino-1-deoxy-beta-D-glucopyranosyl)formamide could be isolated. Acetylation of this amino-amide by Ac2O/Py and subsequent debenzoylation gave C-(1-acetamido-1-deoxy-beta-D-glucopyranosyl)formamide. Applying the same conditions to the crude reduction mixture allowed the alpha-anomer to be isolated as a minor component. An alternative pathway to produce the above beta-anomer appeared in the reaction of C-(1-bromo-1-deoxy-beta-D-glucopyranosyl)formamide with CH3CN in the presence of Ag2CO3 to yield 1-acetamido-2,3,4,6,-tetra-O-benzoyl-1-deoxy-beta-D-glucopyranosyl cyanide, which was hydrated, in the presence of TiCl4, to the formamide. Some of the new compounds were shown to be weak inhibitors of muscle glycogen phosphorylase b.

Enzyme Inhibitors↗
PubMed1966

Components of the cell wall of Clostridium welchii (type A).

1. The cell wall of Clostridium welchii (type A) contains alanine, 2,6-diaminopimelic acid, glutamic acid, glycine, glucosamine, muramic acid, galactosamine, mannosamine, ethanolamine, rhamnose, galactose and phosphorus. 2. Heating with formamide at 150 degrees resolved the wall into a formamide-soluble polysaccharide fraction and a formamide-insoluble mucopeptide fraction. 3. The formamide-soluble fraction contained two components: an electrophoretically neutral polysaccharide made up of galactose, rhamnose, galactosamine and phosphorus and an electrophoretically acidic polymer containing mannosamine, ethanolamine and phosphorus. 4. The formamide-insoluble residue has been digested by lysozyme to give soluble fragments of high molecular weight. 5. All fractions contain an unknown ethyl acetate-extractable substance that can be oxidized by sodium metaperiodate. 6. The amino acid compositions of the fragments produced by lysozyme are compatible with a mucopeptide structure which has cross bridges containing all of the constituent amino acids.

Amino Acids↗
PubMed1996

Reevaluation of the exact CAG repeat length in hereditary cerebellar ataxias using highly denaturing conditions and long PCR.

Hereditary cerebellar ataxias, including spinocerebellar ataxia type I (SCA1), dentato-rubro-pallidoluysian atrophy (DRPLA), and Machado-Joseph disease (MJD), have been associated with unstable CAG repeats. The length of the CAG repeat is a major factor in determining the age of onset of these diseases. In electrophoresis through acrylamide gels with formamide, the CAG repeat length following the polymerase chain reaction (PCR) coincides with the sequence-determined repeat length after subcloning. However, without formamide, PCR products with long CAG repeats appear 1-4 repeats shorter than when electrophoresed with formamide, and the repeat lengths are variable. In addition, the larger the CAG repeats are, the more difficult are the PCR reactions. A mixture containing thermostable Taq and Pwo DNA polymerases (so-called "long PCR") is much more sensitive than that with Taq polymerase alone in detecting- expanded CAG repeats. Therefore, highly denaturing conditions, especially formamide gel electrophoresis, and the "long PCR" protocol should be used to evaluate the exact CAG repeat length. We have used these principles to detect unstable CAG repeats. The normal ranges are 14-34 repeats for MJD, 6-31 repeats for DRPLA, and 21-32 repeats for SCA1.

Alleles↗
PubMed1989

Optimization of in situ hybridization to human metaphase chromosomes.

A simplified method describing optimal conditions for in situ hybridization to human chromosomes is presented. A 1.5-kb DNA fragment coding for part of the 28 S rRNA was subcloned into pSP65. Tritium-labeled RNA was synthesized as runoff transcripts and 39-67% of the labeled probes specifically hybridized to the nucleolar organizer regions of the acrocentric chromosomes. Denaturation performed with 70% formamide, 1 mM EDTA, 2 X SSC gave a high specific hybridization with both fresh chromosome spreads (1-8 weeks) and older preparations (3-6 months). To obtain good chromosome morphology and a high specific hybridization it was important to neutralize the final formamide denaturation mixture containing 70% formamide, 1 mM EDTA, and 2 X SSC, whereas it was unimportant to deionize the formamide. Freshly made slides denatured with 0.15 M NaOH in 70% ethanol hybridized equally well with the rRNA probe. Despite treatment of the chromosomes with RNases before denaturation the following proteinase K and the acetylation steps recommended could be omitted without degradation of the rRNA probe as judged from the high specific hybridization to the nucleolar organizer regions.

Animals↗
PubMed2006

The effect of solvent polarity on the molecular surface properties and adhesion of Escherichia coli.

The elasticity and molecular surface characteristics of Escherichia coli JM109 were investigated via atomic force microscopy (AFM) in solvents expressing different polarities. The nature of bacterial adhesion and surface characteristics was probed in formamide, water, and methanol, with dielectric constants of 111, 80, and 33, respectively. Solvent polarity affected the elasticity of the bacterium, the conformation of the cell surface biopolymers, the height of the surface biopolymers, and measured adhesion forces between the bacterium and silicon nitride. By applying the Hertz model to force-indentation data, we determined that the Young's modulus was greatest in the least polar solvent, with values of 182 +/- 34.6, 12.8 +/- 0.1, and 0.8 +/- 0.3 MPa in methanol, water, and formamide, respectively. The thickness of the biopolymer brush layer on the bacterial surface was quantified using a steric model, and these values increased as polarity increased, with values of 27, 93, and 257 nm in methanol, water, and formamide, respectively. The latter results suggest that highly polar conditions favor extension of the biopolymer brush layer. Cross-sectional analysis performed on tapping mode images of the bacterial cells in methanol, water, and formamide further supported this hypothesis. The image height values are larger, since the image analysis measures the height of the bacterium and the polymer layer, but the trend with respect to solvent polarity was the same as was obtained from the steric model of the brush length. Measured adhesion forces scaled inversely with solvent polarity, with greatest adhesion observed in the least polar solvent, methanol. The combined conformational changes to the bacterial surface and biopolymer layer result in different presentations of macromolecules to a substrate surface, and therefore affect the adhesion forces between the bacterial molecules and the substrate. These results suggest that polarity of the solvent environment can be manipulated as a design parameter to control or modify the bacterial adhesion process.

Bacterial Adhesion↗
PubMed1998

The water channel of cytochrome c oxidase: inferences from inhibitor studies.

Cytochrome c oxidase couples electron transfer to proton transfer from inside the mitochondrion to the cytosol. Protons pass through a channel; it is closed except when protons are pumped. Electron transfer is also coupled to a water cycle. Water moves into and out of the oxidase during electron transfer, presumably through a channel. The three processes are coupled because of the common dependence on electron transfer. If water and protons had to pass through the same channel for the proton to pass, it might be possible to block the pore by entraining small molecules in the flow. The data in this report indicate that there is a correlation between the ability of a compound to inhibit the oxidase and its size. Formamide and formaldehyde are potent inhibitors. Larger and smaller molecules are poor inhibitors. Formamide introduces an internal block in electron transfer. It is a slow-onset, reversible inhibitor, dependent on turnover to manifest its effects. Vesicular oxidase is less influenced by formamide than is soluble oxidase; formamide must pass a permeability barrier to act. The data are consistent with a proton channel with constrictions at both ends that open to yield a pore of approximately 4 A.

Animals↗
PubMed2005

Spontaneous formation of gold nanoparticles in poly(ethylene oxide)-poly(propylene oxide) solutions: solvent quality and polymer structure effects.

We report here on the effects that the solution properties of poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) block copolymers have on the reduction of hydrogen tetrachloroaurate(III) hydrate (HAuCl4.3H2O) and the size of gold nanoparticles produced. The amphiphilic block copolymer solution properties were modulated by varying the temperature and solvent quality (water, formamide, and their mixtures). We identified two main factors, (i) block copolymer conformation or structure (e.g., loops vs entanglements, nonassociated polymers vs micelles) and (ii) interactions between AuCl4- ions and block copolymers (attractive ion-dipole interactions vs repulsive interactions due to hydrophobicity), to be important for controlling the competition between the reactivities of AuCl4- reduction in the bulk solution to form gold seeds and on the surface of gold seeds (particles) and the particle size determination. The particle size increase observed with increased temperature in aqueous solutions is attributed to enhanced hydrophobicity of the block copolymer, which favors AuCl4- reduction on the surface of seeds. The lower reactivity and higher particle sizes observed in formamide solutions are attributed to the shielding of ion-dipole interaction between AuCl4- ions and block copolymers by formamide, which overcomes the beneficial effects of formamide on the block copolymer conformation (lower micelle concentration).

Chlorides↗
PubMed1992

Utilization of cyanide as nitrogenous substrate by Pseudomonas fluorescens NCIMB 11764: evidence for multiple pathways of metabolic conversion.

The growth of Pseudomonas fluorescens NCIMB 11764 on cyanide as the sole nitrogen source was accomplished by use of a modified fed-batch cultivation procedure. Previous studies showing that cyanide metabolism in this organism is both an oxygen-dependent and an inducible process, with CO2 and ammonia representing conversion products, were confirmed. However, washed cells (40 mg ml-1 [dry weight]) metabolized cyanide at concentrations far exceeding those previously described; 85% of 50 mM KCN was degraded in 6 h. In addition, two other C1 metabolites were detected in incubation mixtures; their identities were confirmed as formamide and formate by 13C nuclear magnetic resonance spectrocopy, high-pressure liquid chromatography, radioisotopic trapping experiments, and other analytical means. The relative yields of all four metabolites (CO2, formamide, formate, and ammonia) were shown to be dependent on the KCN concentration and availability of oxygen; at 0.5 to 10 mM substrate, CO2 was the major C1 product, whereas at 20 and 50 mM substrate, formamide and formate were principally formed. The latter two metabolites also accumulated during prolonged anaerobic incubation, suggesting that P. fluorescens NCIMB 11764 can elaborate several pathways of cyanide conversion. One is formally similar to that proposed previously (R. E. Harris and C. J. Knowles, FEMS Microbiol. Lett. 20:337-341, 1983), involving the oxygen-dependent conversion of cyanide to CO2 and ammonia. The other two, occurring in the presence or absence of oxygen, involve separate reactions to yield, respectively, formate plus ammonia or formamide.(ABSTRACT TRUNCATED AT 250 WORDS)

Ammonia↗
PubMed2002

Single-base-pair discrimination of terminal mismatches by using oligonucleotide microarrays and neural network analyses.

The effects of single-base-pair near-terminal and terminal mismatches on the dissociation temperature (T(d)) and signal intensity of short DNA duplexes were determined by using oligonucleotide microarrays and neural network (NN) analyses. Two perfect-match probes and 29 probes having a single-base-pair mismatch at positions 1 to 5 from the 5' terminus of the probe were designed to target one of two short sequences representing 16S rRNA. Nonequilibrium dissociation rates (i.e., melting profiles) of all probe-target duplexes were determined simultaneously. Analysis of variance revealed that position of the mismatch, type of mismatch, and formamide concentration significantly affected the T(d) and signal intensity. Increasing the concentration of formamide in the washing buffer decreased the T(d) and signal intensity, and it decreased the variability of the signal. Although T(d)s of probe-target duplexes with mismatches in the first or second position were not significantly different from one another, duplexes with mismatches in the third to fifth positions had significantly lower T(d)s than those with mismatches in the first or second position. The trained NNs predicted the T(d) with high accuracies (R(2) = 0.93). However, the NNs predicted the signal intensity only moderately accurately (R(2) = 0.67), presumably due to increased noise in the signal intensity at low formamide concentrations. Sensitivity analysis revealed that the concentration of formamide explained most (75%) of the variability in T(d)s, followed by position of the mismatch (19%) and type of mismatch (6%). The results suggest that position of the mismatch at or near the 5' terminus plays a greater role in determining the T(d) and signal intensity of duplexes than the type of mismatch.

Base Pair Mismatch↗
PubMed1975

Antigens of Streptococcus mutans: characterization of a polysaccharide antigen from walls of strain GS-5.

A cell wall-associated polysaccharide antigen was isolated from Streptococcus mutans GS-5 and appeared to determine serotype c specificity. Ouchterlony double-diffusion analysis of crude formamide extracts derived from purified cell walls of two serotype c strains (GS-5 and JC-2) showed complete identify when reacted with anti-GS-5 sera. Immunoelectrophoresis of this extract demonstrated the typical mobility for this serotype as described by others. Column chromatography on BioGel P-100 of the crude formamide extracts derived from GS-5 walls resulted in a single antigenic peak being resolved. This material, when loaded onto a diethylaminoethylcellulose column and eluted with a linear gradient of ammonium carbonate (0.0 to 0.2 M), was resolved further into two serologically reactive peaks (I and II). Only two consituents, rhamnose and glucose, were detected in the purified column fractions. Peak 1 had a rhamnoseto-glucose molar ratio of 0.9:1.0, and peak II, the major resolvable fraction, had a molar ratio of 1.7:1.0, The peak II ratio was very similar to that found in the formamide extract residue pellet (1.6:1.0)9 Ouchterlony analysis of the crude formamide extract and the purified fractions revealed only partial identify between peaks I and II but complete identify between peak II and the crude extract. Likewise, immunoelectrophoresis showed no differences in mobility of peak II and the crude extract, whereas peak I moved towards the cathode. Possible structural relationships between the two antigenic fractions are discussed below. Hapten inhibition studies suggested that an alpha-glucosyl group is at the immunodeterminant site of the antigen.

Antigens, Bacterial↗
PubMed1977

Messenger ribonucleoprotein complexes isolated by oligodeoxythymidylate-cellulose chromatography from Neurospora crassa polysomes.

Messenger ribonucleoprotein (mRNP) complexes were isolated from ethylenediaminetetraacetic acid-dissociated polysomes of Neurospora crassa. Approximately 15% of the [3H]uridine incorporated into polysomal ribonucleic acid (RNA) during a 15-min pulse was eluted from oligodeoxythymidylate-cellulose as an mRNP complex. The isolated mRNP complexes exhibited sedimentation coefficients ranging from 15S to greater than 60S. RNA isolated from these mRNP complexes sedimented in sucrose gradients between 4S and 40S, with broad peaks at 15S and 24S. The buoyant density of mRNP complexes eluted with 25% formamide was 1.42 to 1.44 g/cm3, whereas for mRNP complexes eluted with 50% formamide it was 1.48 to 1.50 g/cm3. Six polypeptides, with molecular weights of 14,000, 19,000, 24,000, 31,000, 44,000, and 66,000, were associated with mRNP complexes eluted with 25% formamide. The mRNP complexes eluted with 50% formamide had one associated polypeptide, of molecular weight 27,000.

Centrifugation, Density Gradient↗
PubMed2002

Solvation of dried dentin matrix by water and other polar solvents.

PURPOSE: To develop a simple method for measuring the degree of solvation of dried, demineralized dentin matrix by water and other polar solvents. The null hypothesis was that there are no differences in expansion forces produced by different polar solvents. MATERIALS AND METHODS: Midcoronal dentin discs were prepared from extracted, unerupted human third molars. The discs were cut into square specimens with surface areas of 2 x 2, 3 x 3 and 4 x 4 mm and thicknesses of 0.5, 1.0 and 1.5 mm. After demineralization in 0.5 M EDTA (pH 7), the dimensions of the specimens were measured both wet and dry. Dry specimens were held between two parallel steel plates connected to a 50 N load cell which measured the solvation force when water or other polar solvents were added. After measuring the expansion force induced by water, the specimens were fixed in glutaraldehyde and the trials repeated. On additional specimens, repeated measures of expansion forces were obtained using water, methanol, ethanol, n-propanol, n-butanol, ethylene glycol, formamide, hydroxyethylmethacrylate, N,N-dimethyl formamide and acetone in unfixed specimens. RESULTS: Water produced hydration forces as high as 204 g before, and 428 g after glutaraldehyde treatment. The hydration force correlated better with specimen thickness than with surface area. Water solvated the matrix faster than methanol > ethanol > formamide > ethylene glycol. Hydroxyethylmethacrylate, N,N-dimethyl formamide and acetone were unable to solvate the dried matrix. Regression analysis of solvation force vs. Hansen's solubility parameters for dispersive, polar and hydrogen bonding forces demonstrated that solvation force correlations were highest with hydrogen bonding solubility parameters. Measurements of solvation forces provides a simple method for determining solvent-collagen matrix interactions.

1-Butanol↗
PubMedDate not supplied

[The effect of unithiol on the antineoplastic and antimetastatic activity of N-methylformamide].

Antitumor and antimetastatic properties of N-methyl formamide--an agent related to differentiation of tumoral cells and unithiol, official detoxicating drug containing SH-groups were studied in C57Bl/6 mice inoculated with Lewis's lung carcinoma. The drugs were administered intraperitoneally into animals at days 1, 4, 7, 13 and 16 after carcinoma inoculation and their effects were evaluated at day 19. Single administration of unithiol, 5 mg/kg body weight, did not affect tumor growth and metastases spreading. At the same time, N-methyl formamide (single administration of 300 mg/kg body weight) inhibited tumor growth and decreased 5-fold an amount of spontaneous metastases in lungs. However, the antitumor and antimetastatic activities of N-methyl formamide were decreased after simultaneous administration with unithiol. Importance of SH-groups in therapeutic effects of N-methyl formamide is discussed.

Animals↗
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↗