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PubMed1990

Ultraviolet imaging densitometry of unstained gels for two-dimensional electrophoresis.

A system suitable for ultraviolet imaging densitometry of two-dimensional electrophoretic gels that are unstained is described, together with its applications. A flying-spot densitometer linked with a personal computer was used for data acquisition, generation of mapping data, and image processing. Randomly distributed zones of proteins on two-dimensional gels were detected at 280 nm without being stained by two-dimensional scanning, and the densitometric value of each pixel (0.2 x 0.2 mm) was memorized by the computer, which generated a mapping pattern with density contours. The amount and densitometric value of cytochrome c had a linear relationship in the range of 2-200 micrograms. Zone locations of bovine liver proteins separated on two-dimensional gels were indicated on a map expressed in X-Y coordinates, and the pIs and molecular weights could be calculated from the map by use of pI and molecular weight markers on the same gel.

Animals↗
PubMed1989

Mass determination of receptor-mediated accumulation of phosphatidate and diglycerides in human neutrophils measured by Coomassie blue staining and densitometry.

Quantitation of 1,2-diacylglycerol (AAG), 1-0-alkyl-2-acylglycerol (EAG) and phosphatidic acid (PA) was conducted in polymorphonuclear leukocytes (PMN) labeled with 1-0-[3H]alkyl-2-acyl-GPC following stimulation with 1 microM fMLP using Coomassie blue staining and densitometry. At 5s AAG and PA increased by 80% and 107%, respectively, over controls. The accumulation of PA, which reached a maximum by 30s, was higher than AAG by 302% at 5s, and 550% at 30s. EAG accumulation was delayed by 15s following stimulation of PMN. These results show that AAG accumulates before EAG and support the role of AAG in cellular activation, perhaps, via the stimulation of protein kinase C (PKC). EAG may serve to counter the effects of AAG or may itself elicit responses. The high concentrations of PA which accumulate early suggest that PA may be generated by the activation of phospholipase D in PMN stimulated with fMLP.

Chromatography, Thin Layer↗
PubMed1994

Monitoring of monoclonal gammopathies: rational use of densitometry and rate nephelometry.

The follow-up of patients with monoclonal gammopathies at our institution includes serial serum protein electrophoresis (SPE) with densitometry and IgG, IgA, and IgM quantitative immunoglobulin (QIG) determinations by rate nephelometry. This retrospective audit compares monoclonal protein concentration as estimated by SPE versus QIG in 456 serial serum specimens from 105 patients to determine whether any of the tests provide redundant information. A comparison of the methods demonstrated good correlation between SPE (x-axis) and QIG (y-axis) quantitation for each immunoglobulin class: IgG had a slope of 1.45 and an intercept of 1.60 (Sy/x = 7.46, r = 0.96, n = 250); IgA had a slope of 1.30 and an intercept of -1.37 (Sy/x = 6.85, r = 0.96, n = 78); and IgM had a slope of 1.95 and an intercept of 2.06 (Sy/x = 5.16, r = 0.98, n = 128). The data for individual patients showed similar good correlations. Exceptions included IgA peaks "buried" in the beta region of the SPE (resulting in invalid SPE estimates of monoclonal protein concentration), and IgG peaks of less than 10 g/L (when background polyclonal IgG immunoglobulin skews the QIG estimate of monoclonal protein concentration). An algorithm is proposed whereby monoclonal protein concentration is measured by the specific QIG (i.e., IgG, IgA, or IgM) determination for the routine monitoring of patients, except for those with IgG peaks of less than 10 g/L that are followed by SPE.

Blood Protein Electrophoresis↗
PubMed1995

Do porous calcium hydroxyapatite ceramics cause porosis in bone? A bone densitometry and biomechanical study on cortical bones of rabbits.

Porous calcium hydroxyapatite (CHA) ceramics are biocompatible and present osteoconductive properties. These ceramics are widely used in orthopaedic surgery; however, it is not yet known whether they have some adverse effects on bone and bone marrow healing. Our previous radiological study revealed possible local porosis at the adjacent sites of the CHA ceramic. Histological findings of the same study revealed bone marrow swelling and depletion at the implantation site. Osteoclasts removed particles of the implant that may be the cause of local porosis. In the present study, possible local osteoporosis was evaluated by bone densitometry analyses, and compression and three-point bending tests. CHA particles were implanted into the left limbs and a sham operation was utilized on the right limbs of 75 white rabbits. The animals were followed up for 23 weeks for bone mineral density and for 6 months for biomechanical analyses. The CHA implanted area and its distal or proximal adjacent areas were evaluated with a Hologic QDR-2000 bone densitometer. Three-point bending and compression tests were performed with an M-30 K material testing device. The results revealed a time-dependent bone density increase at the CHA implantation site and no significant porosis at adjacent areas of the implant. The stiffness of CHA-implanted bones in three-point bending is larger than that of the control group. CHA-implanted rabbit bones presented a different fracture pattern from the control group. The stiffness of the control and CHA-implanted bones generally increased with time indicating no adverse effects of porous CHA ceramics in bone and bone marrow healing. The clinical relevance of this work is that porous CHA ceramics do not cause local porosis at adjacent areas when implanted into osseous sites.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Determination of plasmid copy number by fluorescence densitometry.

A simple and reliable method for the determination of plasmid copy numbers by direct fluorescence densitometry of ethidium bromide-stained electrophoretic gels was developed. In developing the method, the following parameters were evaluated and controlled: plasmid DNA trapping in the linear chromosomal DNA, staining-destaining kinetics for ethidium bromide, linearity of the fluorescence response, and the effect of the molecular topology of DNA on ethidium bromide binding to DNA in agarose.

DNA, Bacterial↗
PubMed1985

Quantification of ethanol-induced gastric mucosal injury by transmission densitometry.

A method to objectively quantify the extent of ethanol-induced gastric lesions has been developed. The method utilizes a transmission densitometer to measure the optical density of the photographic negative of the stomach mucosa. Tissues from ethanol- and untreated animals are compared with tissues from animals pretreated with prostanoids before ethanol; this method permits a reproducible and objective evaluation of mucosal protection. We demonstrate that the optical density is proportional to subjective score and damage, and that the densitometry method differentiates between the protective effect of different doses of 16,16-dimethyl prostaglandin E2 and natural prostaglandin E2.

Animals↗
PubMed1984

Video and scanning microdensitometer-based imaging systems in autoradiographic densitometry.

The characteristics of computer-assisted imaging devices (CIDs) incorporating charge-coupled device cameras, vidicon cameras and scanning microdensitometers (SMDs) are compared. Advantages of vidicons include their low cost, the option of real-time imaging, continuously variable optical magnification and the wide choice of available vidicon elements. Disadvantages include densitometric non-linearities and, with some vidicons, lack of dynamic range. A CID comprising a laboratory minicomputer, image processing device, vidicon and SMD scanners and our 'PANDA' image analysis routines is described. Software corrections for vidicon errors are evaluated by comparing the densitometric accuracy of the CID receiving input from the vidicon camera and from the SMD. The scanners exhibit similar densitometric accuracy and linearity, and acceptable between-session repeatability. It is concluded that densitometric precision of a corrected vidicon scanner is not the limiting factor for quantitative autoradiographic densitometry.

Animals↗
PubMed1988

Combined thin-layer chromatography-photography-densitometry for the quantitation of cyclophosphamide and its four principal urinary metabolites.

A novel method for the quantitative determination of the anti-cancer drug cyclophosphamide and its principal urinary metabolites 4-oxocyclophosphamide, carboxyphosphamide, phosphoramide mustard and bis(2-chloroethyl)amine has been devised. The assay combines adsorption of drug-related material onto Amberlite XAD-2 and thin-layer chromatography with spot visualization using 4-(4-nitrobenzyl)pyridine, rapid photography and densitometry. The intra-assay coefficient of variation for each compound was less than 6%. The limit of detection of the assay was 1 microgram ml-1 for cyclophosphamide, phosphoramide mustard and bis(2-chloroethyl)amine and 0.5 microgram ml-1 for 4-oxocyclophosphamide and carboxyphosphamide. The method was validated for cyclophosphamide and 4-oxocyclophosphamide using gas chromatography. It is concluded that the method provides the first means of determining the full metabolic spectrum for cyclophosphamide in patients without recourse to the administration of radioisotopically labelled drug.

Biotransformation↗
PubMed1991

Densitometry for analysis of protein and peptide hydrolysates: application to luteinizing hormone releasing hormone.

Densitometric analysis has been performed on various standard protein and peptide solutions and on luteinizing hormone releasing hormone hydrolysate. After thin-layer chromatographic separation using two-dimensional separation on cellulose plates, the plates were mapped and each amino acid was assayed individually in order to obtain a specific integration profile of the hydrolysate. Densitometry proved to be a useful method for the screening of protein hydrolysates and is accurate enough for the assay of peptide hydrolysates. The hormone was hydrolysed with hydrochloric acid-trifluoroacetic acid (1:1, v/v) in the presence of thioglycolic acid. This limits the loss of tryptophan and does not interfere with the analysis. Results were sufficiently reliable, accurate and reproducible for routine analysis.

Chromatography, High Pressure Liquid↗
PubMed1993

Analysis of complex mixtures of phospholipid classes from cell membranes using two-dimensional thin-layer chromatography and scanning laser densitometry.

Increasing recognition of the important roles served by membrane phospholipids in cellular metabolic and signal transduction processes has stimulated interest in examining potential phospholipid abnormalities in patients with psychiatric disorders. This report describes a method, based on several novel modifications of existing techniques, for concurrently analyzing nanomolar amounts of nine phospholipid classes in a single aliquot of membrane extract. With this method, diverse phospholipid classes are first separated by two-dimensional thin-layer chromatography, and then determined using two-dimensional scanning laser densitometry. The method is able to quantitate even small amounts of specific phospholipid classes, corresponding to < 10 ng of lipid phosphorus. The sensitivity of this method allows it to be readily applied to clinical studies involving membranes from cell types that are obtainable only in small quantities.

Blood Platelets↗
PubMed1993

Simplified method for simultaneous determination of diazepam and its metabolites in urine by thin-layer chromatography and direct densitometry.

A direct densitometric method for determination of diazepam and its metabolites in urine was developed. The proposed procedure involves acid hydrolysis of urine specimens, thereby converting diazepam and its metabolites into benzophenones [2-methylamino-5-chlorobenzophenone (MACB) and 2-amino-5-chlorobenzophenone (ACB)]. It is followed by extraction with chloroform-isopropanol (3:1, v/v). The two benzophenones were separated on thin-layer chromatography plates using hexane-diethyl ether-acetic acid (80:10:10) as a mobile phase. Quantitation of the MACB and ACB spots was achieved by direct ultraviolet densitometry. The limit of detection was 0.5 micrograms per ml of urine for both benzophenones. The proposed method is simple, rapid, reproducible and has been found to be effective for direct determination of diazepam and its metabolites in urine.

Benzophenones↗
PubMed1989

Determination of thiols of biological and pharmacological interest by high-performance thin-layer chromatography and fluorescence scanning densitometry.

The application of high-performance thin-layer chromatography (HPTLC) with fluorescence scanning densitometry provides a simple, rapid and reliable system for the qualitative and quantitative determination of several thiols of biological and pharmacological interest. The determination of a mixture of thiols (captopril, coenzyme A, cysteamine, cysteine and glutathione), together with their disulphides may readily be performed by pre-chromatographic derivatization with the thiol-specific fluorobenzoxadiazole reagents SBD-F and ABD-F, followed by HPTLC separation on silica gel plates using isopropyl ether-methanol-water-acetic acid (9:8:2:1, v/v/v/v) as the developing solvent, and fluorodensitometric measurement of the fluorescing derivatives. Detection limits of about 30 pg (coenzyme A) to 6 pg (cysteamine) per spot were achieved; the relative standard deviation (RSD) of the complete procedure was 1.16-3.2%.

Acetates↗
PubMedDate not supplied

Analysis of FCE 23762 (methoxymorpholinodoxorubicin hydrochloride), a new antitumour agent, by HPTLC and scanning densitometry.

A simple, rapid and reproducible high-performance thin-layer chromatographic (HPTLC) method using UV or fluorescence scanning densitometry has been developed for the assay and purity control of methoxymorpholinodoxorubicin hydrochloride (FCE 23762). With a mobile phase of chloroform-methanol-acetic acid (93:6:1, v/v/v) and a silica gel plate, all potential impurities were separated from the main component and from each other. Detection limits at a signal-to-noise ratio of 2:1 were a few nanograms for UV detection and < 1 ng for fluorescence emission. The RSD values for the examined compounds were all < 3%.

Acetates↗
PubMed1995

HPTLC and reflectance mode densitometry of anthocyanins in Malva silvestris L.: a comparison with gradient-elution reversed-phase HPLC.

Aqueous alcoholic mallow flower extracts were analyzed both by HPTLC-densitometry in the reflectance mode at 530 nm and by reversed-phase HPLC with gradient elution. For the mallow flower anthocyanins the best chromatographic resolution was obtained by HPLC, which revealed only two main compounds, confirmed by FAB-MS: malvidin 3,5-O-diglucoside (malvin) and malvidin 3-O-(6"-O-malonylglucoside)-5-O-glucoside. The HPTLC densitometric method on cellulose plates provides accuracy, reproducibility and selectivity for the quantitative analysis of the anthocyanins and this method was shown to be much more sensitive than the HPLC-DAD system, at 530 nm. Both methods give comparable quantitative results for total anthocyanins when applied to mallow flowers from two different sources: Italy and Albania.

Anthocyanins↗
PubMedDate not supplied

A possible way to solve problems in CT densitometry.

The common computed tomography (CT) densitometric scale expressed in Hounsfield units does not possess sufficient representativity. A standard scale is needed to solve problems in CT densitometry. A new scale based on a special phantom with different solutions of calcium chloride permits the elimination of shortcomings of the common scale. Indications for testing CT scanners with the help of the phantom are offered.

Densitometry↗
PubMed2004

Assessment of local changes of cerebral perfusion and blood concentration by near infrared spectroscopy and ultrasound contrast densitometry.

The objective of this study is to correlate regional cerebral blood concentration measurements made with near infrared spectroscopy to simultaneous local measurements of ultrasound contrast agent (CA) densitometry. Experiments were performed with piglets (7 kg) under general anesthesia. The cerebral blood flow (CBF) and volume (CBV) were changed by inducing various degrees of hypercapnia. NIRS measurements were performed with a quasi-continuous wave system, using an optode distance of 3-6 cm. The concentration changes in oxygenated and deoxygenated hemoglobin and their sum and difference (cO2Hb, cHHb, ctHb, cHbD) were continuously calculated. Ultrasound contrast agent (SF6) was administered as a short intra-venous bolus. Ultrasound equipment was used in pulse inversion second harmonic gray scale imaging mode at low transmit power setting. Three regions-of-interest (0.25 cm2) were analyzed in each image. Wash-in curves were constructed as spatial mean gray level vs. time. The variables collected with both methods changed according to the induced changes in the physiological condition. Changes in the PaCO2, pH and carotid flow induced highly correlated changes in cO2Hb, cHHb, ctHb and cHbD, and in the variables derived from CA analyses. NIRS and CA methods measure regional, respectively, local changes in CBV and CBF. Moreover, NIRS can yield complementary information about the cerebral oxygenation.

Animals↗
PubMed2004

Comparison between HPLC and HPTLC-densitometry for the determination of harpagoside from Harpagophytum procumbens CO(2)-extracts.

Carbon dioxide (CO(2)) extracts of the secondary roots of Harpagophytum procumbens were quantified by high performance liquid chromatography (HPLC) and high performance thin layer chromatography (HPTLC). An isocratic HPLC method was used for the quantification of the iridoid glucoside harpagoside at 278 nm. A HPTLC assay was developed for the determination of harpagoside after coloration at 509 nm. The diode array detection of both analytical assays were used to examine the purity of harpagoside peaks and compared with the standards, respectively. The assays provide good accuracy, reproducibility and selectivity for the quantitative analysis of harpagoside. The harpagoside contents of 15 different CO(2)-extracts were compared by HPLC and HPTLC-densitometry. The quantitative results of both analytical methods did not show any statistical significance between each other, although a trend to slightly lower mean values could be found for the HPTLC method.

Analgesics↗
PubMed2006

Determination of phenothiazine derivatives by high performance thin-layer chromatography combined with densitometry.

A high performance thin-layer chromatography (HPTLC) method combined with densitometry for determination of phenothiazine derivatives is described. Quantitation was performed in reflectance mode by using a computer-controlled densitometer Desaga CD 60. Established calibration curve (r > 0.999), precision (RDS values: 0.95-2.53%), detection limits as well as recovery values (101.1-102.8%) were found to be satisfactory. The presented method is rapid, precise and sensitive, and may be alternative to traditionally used HPLC. The method has been successfully applied in the analysis of pharmaceutical formulations.

Antiprotozoal Agents↗