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PubMed1996

'Mutational SURF': a strategy for improving lead compounds identified from combinatorial libraries.

Synthesis and testing of mixtures of compounds in a combinatorial library offers the potential of much greater throughput than the 'one compound, one well' approach. When mixtures of compounds are screened, however, pooling and deconvolution strategies must be employed to identify the most active compound in the library. The possibility exists that the most active compound will not be identified. We have developed a theoretical model of library deconvolution using the well characterized properties of nucleic acid hybridization to calculate activities of individual molecules in libraries of more than 250,000 compounds. Calculations using this model have been employed to evaluate strategies for pooling and deconvolution. In the presence of errors in synthesis and testing, iterative deconvolution or position scanning sometimes identified a compound with sub-optimal activity. We describe a procedure called 'mutational SURF' in which 'mutants' of the selected compound are individually synthesized and tested. Simulations of mutational SURF using our model libraries suggest that mutational SURF provides an efficient method for improving the activity of lead compounds identified from combinatorial libraries.

Base Sequence↗
PubMed2004

Trilinear chemometric analysis of two-dimensional comprehensive gas chromatography-time-of-flight mass spectrometry data.

Two-dimensional comprehensive gas chromatography (GC x GC) is a powerful instrumental tool in its own right that can be used to analyze complex mixtures, generating selective data that is applicable to multivariate quantitative analysis and pattern recognition. It has been recently demonstrated that by coupling GC x GC to time-of-flight mass spectrometry (TOFMS), a highly selective technique is produced. One separation on a GC x GC/TOFMS provides retention times on two chromatographic columns and a complete mass spectrum for each component within the mixture. In this manuscript, we demonstrate how the selectivity of GC x GC/TOFMS combined with trilinear chemometric techniques such as trilinear decomposition (TLD) and parallel factor analysis (PARAFAC) results in a powerful analytical methodology. Using TLD and PARAFAC, partially resolved components in complex mixtures can be deconvoluted and identified using only one data set without requiring either signal shape assumptions or fully selective mass signals. Specifically, a region of overlapped peaks in a complex environmental sample was mathematically resolved with TLD and PARAFAC to demonstrate the utility of these techniques as applied to GC x GC/TOFMS data of a complex mixture. For this data, it was determined that PARAFAC initiated by TLD performed a better deconvolution than TLD alone. After deconvolution, mass spectral profiles were then matched to library spectra for identification. A standard addition analysis was performed on one of the deconvoluted analytes to demonstrate the utility of TLD-initiated PARAFAC for quantification without the need for accurate retention time alignment between sample and standard data sets.

Gas Chromatography-Mass Spectrometry↗
PubMed2000

Fine-structure enhancement--assessment of a simple method to resolve overlapping bands in spectra.

A simple mathematical procedure--fine-structure enhancement--has been assessed on its ability to resolve overlapping bands in spectra. Its advantages and limitations have been explored using synthetic and experimental spectra. Fine-structure enhancement involves smoothing the original spectrum, multiplying the smoothed spectrum with a weighting factor and subtracting this spectrum from the original spectrum. As a result, the fine-structure of the original spectrum is enhanced in the processed spectrum and bands that overlap in the original spectrum appear as distinct bands in the processed spectrum. To be resolved by fine-structure enhancement, Lorentzian lines have to be separated by more than their quarter width at half maximum, Gaussian lines by more than their half width at half maximum. A comparison of fine-structure enhancement and Fourier self-deconvolution shows that Fourier self-deconvolution has in theory a higher potential to resolve overlapping bands. However, this depends crucially on the correct choice of the parameters. In practice, when parameters commonly used are chosen for Fourier self-deconvolution, fine-structure enhancement leads to similar results. This is demonstrated at the example of the infrared absorbance spectrum of the protein papain, where the amide I band components could be resolved similarly with both methods. Thus, fine-structure enhancement seems to be a simple alternative to Fourier self-deconvolution that does not require specialised software.

Models, Chemical↗
PubMed2004

A strategy for identifying differences in large series of metabolomic samples analyzed by GC/MS.

In metabolomics, the purpose is to identify and quantify all the metabolites in a biological system. Combined gas chromatography and mass spectrometry (GC/MS) is one of the most commonly used techniques in metabolomics together with 1H NMR, and it has been shown that more than 300 compounds can be distinguished with GC/MS after deconvolution of overlapping peaks. To avoid having to deconvolute all analyzed samples prior to multivariate analysis of the data, we have developed a strategy for rapid comparison of nonprocessed MS data files. The method includes baseline correction, alignment, time window determinations, alternating regression, PLS-DA, and identification of retention time windows in the chromatograms that explain the differences between the samples. Use of alternating regression also gives interpretable loadings, which retain the information provided by m/z values that vary between the samples in each retention time window. The method has been applied to plant extracts derived from leaves of different developmental stages and plants subjected to small changes in day length. The data show that the new method can detect differences between the samples and that it gives results comparable to those obtained when deconvolution is applied prior to the multivariate analysis. We suggest that this method can be used for rapid comparison of large sets of GC/MS data, thereby applying time-consuming deconvolution only to parts of the chromatograms that contribute to explain the differences between the samples.

Algorithms↗
PubMed1999

On-bead combinatorial approach to the design of chiral stationary phases for HPLC.

A library of 36 L-amino acid anilides, which are potential selectors for chiral HPLC, was synthesized in solution and attached to functionalized macroporous polymer beads. The best selector from the library was identified by a deconvolution process using the HPLC separation of several racemic N-(3,5-dinitrobenzoyl)-alpha-amino acid alkylamides as a probe. In each deconvolution step, a series of chiral stationary phases (CSPs) containing a subset of the amino acid anilide selector library was screened for enantioselectivity. After the best CSP was chosen, the library was further deconvoluted until the single best selector was found. The highest selectivity was obtained with a L-proline-1-indananilide that exhibited alpha values up to 23 under normal-phase HPLC conditions. In addition, six CSPs were prepared using individual selectors from the library, and screening results indicate that the deconvolution process indeed led to the most selective receptor.

Amino Acids↗
PubMed1995

A method for processing diffraction data from twinned protein crystals and its application in the structure determination of an FAD/NADH-binding fragment of nitrate reductase.

A general method to deconvolute oscillation data sets from twinned protein crystals to a corresponding single-crystal data set has been developed and applied to diffraction data measured from crystals of a fragment containing the FAD- and NADH-binding domains of nitrate reductase. The procedure allows straightforward processing of diffraction data from twinned crystals. Typically, R(merge) values of reduced data sets from the nitrate reductase crystals after deconvolution are about 0.06 compared to 0.13 and higher before deconvolution. Based on these deconvoluted data sets, the structure of the FAD- and NADH-binding domains of nitrate reductase could be solved successfully. The result indicates that crystal twinning does not necessarily prevent crystallographic structure determination.

Journal Article↗
PubMed1981

Computation of transport function using multiple regression analysis.

A multiple regression analysis, which allows a simultaneous introduction of smoothing, was adopted for a numerical deconvolution process to obtain the transport function from a pair of indicator dilution curves. This process solves concurrently restricting linear quadratic or cubic equations that are fitted to small sections of the transport function curve, an equation that forces the tail part to zero, and uses conditions that restrict the area under the transport function to unity. Each restricting linear equation can be manually stressed by adjusting its corresponding weight iteratively so that unreasonable oscillations emerging on any particular portion of the transport function curve can be suppressed. Stability of the method was tested with various pairs of dilution curves in terms of mathematical models and with those experimentally obtained from the dog portal circulation. The deconvolution by means of the regression analysis with simultaneous smoothing was shown to be far less sensitive to observational errors in the dilution curves or sampling rates than that by means of the direct method without the regression analysis. Multiple regression analysis with simultaneous smoothing was also applied to the deconvolution process using the multiple fitting functions developed by Knopp et al. (Ann. Biomed. Eng. 4: 44-59, 1976). This process was shown to perform a stable deconvolution as well.

Animals↗
PubMed1983

Lateral inverse filtering of ultrasonic B-scan images.

The formation of ultrasonic B-scan images using parallel beams may be modelled as a lateral, one-dimensional convolution of the beam profile and an unknown but wanted reflection coefficient. Lateral inverse filtering, or deconvolution, might therefore be used to improve the image quality. Two different deconvolution techniques are applied to both an image of a tissue mimicking phantom and a human liver. An enhancement of the resolution (defined as the reciprocal of the half-width of the image of a point reflector) of about 1.4 is achieved. This is in good agreement with the previously derived formula R = square root 1n SNR, which relates the signal-to-noise ratio, SNR, to the resolution enhancement, R. However, each method also creates artifacts, and despite the slight resolution enhancement, the deconvoluted liver images do not exhibit more information nor are they more appealing. So it is felt that the computational effort is wasted. This failure is not a fault of the special deconvolution techniques tried here, but rather caused by the logarithmic dependence of R on SNR and by the noise level, which is largely due to macro- and microscopic inhomogeneities of the tissue and cannot be made arbitrarily small.

Filtration↗
PubMed1989

The nature of the gonadotropin-releasing hormone stimulus-luteinizing hormone secretory response of human gonadotrophs in vivo.

To examine the stimulus-secretion response of human pituitary gonadotrophs in vivo, we applied a new multiple parameter deconvolution technique to analyze (1) exogenous GnRH-stimulated LH secretory responses in 10 men with isolated hypogonadotropic hypogonadism (IHH), and (2) endogenous and exogenous GnRH-stimulated LH secretory responses in 8 normal men. The GnRH-deficient men were given 4 bolus doses of synthetic GnRH (7.5, 25, 75, and 250 ng/kg) iv at 2-h intervals in randomized order after long term pulsatile GnRH administration. The normal men were studied by sampling blood at 10-min intervals for 12 h basally and after 2 consecutive 10-micrograms iv GnRH doses. The serum LH peaks in both groups were subjected to quantitative deconvolution to resolve underlying LH secretory and clearance rates simultaneously. Such analyses revealed that exogenous GnRH-induced LH secretory episodes in GnRH-deficient men with IHH could be modeled as algebraically Gaussian distributions of instantaneous LH secretory rates with a mean half-duration of 14 +/- 2 min. The simultaneously resolved half-life of endogenous LH disappearance was 71 +/- 5 min. The log dose-response relationship for GnRH dose vs. maximal LH secretory rate or vs. calculated mass of LH released per secretory burst was linear. In contrast, varying GnRH doses did not alter the duration of LH secretory bursts, the half-time of LH disappearance, or the latency of LH secretory bursts after iv GnRH injections (viz. 7.6 min). Deconvolution analysis of the spontaneous (endogenous GnRH-stimulated) LH peaks in normal men revealed a mean half-duration of secretory bursts of 9.9 +/- 1.5 min, and a mean half-time of endogenous LH disappearance of 76 +/- 5 min. These values were not significantly different from those in the GnRH-treated normal or GnRH-deficient men. In summary, deconvolution analysis of LH release in men with IHH revealed a significant linear relationship between iv doses of pulsed GnRH and computer-resolved LH secretory rate and/or the mass of LH released per secretory event. In contrast, varying doses of GnRH did not alter the lag time between the GnRH stimulus and the LH secretory burst, the duration of LH secretion, or the calculated half-life of the LH released. We conclude that GnRH exerts dose-dependent effects on specific attributes of the secretory response of human gonadotrophs in vivo.

Adult↗
PubMed1984

A clinical evaluation of a prototype positron camera for longitudinal emission tomography.

A multiwire proportional-chamber positron camera, developed at the Rutherford Laboratory, has been evaluated at The Royal Marsden Hospital. The prototype camera consists of two opposing 30 X 30 cm2 chambers. Longitudinal tomograms of a positron-emitting radioactive distribution placed between the detectors are obtained via back-projection and 2D-deconvolution. Due to the limited stereoscopic angle achieved with stationary detectors, only five planes parallel to the detector faces are reconstructed. A selection of images is presented of phantoms using 68Ga and of patients using 18F, 18F fluorodeoxyglucose and Na124I, to illustrate the tomographic performance of the positron camera. A comparison between back-projected and deconvoluted images shows that the 2D-deconvolution process, which includes filtration of image noise, successfully removes the background due to scattered photons. The spatial resolution achieved depends on the half-height frequency cut-off used in the filtering process, and this parameter was chosen according to the count density in the back-projected images. A qualitative visual comparison was made between the positron images and equivalent single photon studies on the same patient. The results justify further development of this new detector, especially for use with generator-produced positron-emitting radionuclides. Three-dimensional deconvolution resulted in improved tomographic performance for phantom data, but was less successful for patient data. These problems associated with limited-angle tomography will be overcome by multi-view acquisition. This study has shown that a relatively low-cost positron imaging system can be used for routine organ imaging. Further developments in hardware and software should yield images which are superior to those from single-photon planar or tomographic studies.

Bone Marrow↗
PubMed2004

MISTRAL: a myopic edge-preserving image restoration method, with application to astronomical adaptive-optics-corrected long-exposure images.

Deconvolution is a necessary tool for the exploitation of a number of imaging instruments. We describe a deconvolution method developed in a Bayesian framework in the context of imaging through turbulence with adaptive optics. This method uses a noise model that accounts for both photonic and detector noises. It additionally contains a regularization term that is appropriate for objects that are a mix of sharp edges and smooth areas. Finally, it reckons with an imperfect knowledge of the point-spread function (PSF) by estimating the PSF jointly with the object under soft constraints rather than blindly (i.e., without constraints). These constraints are designed to embody our knowledge of the PSF. The implementation of this method is called MISTRAL. It is validated by simulations, and its effectiveness is illustrated by deconvolution results on experimental data taken on various adaptive optics systems and telescopes. Some of these deconvolutions have already been used to derive published astrophysical interpretations.

Journal Article↗
PubMed2001

Glucagon-like peptide 1 increases secretory burst mass of pulsatile insulin secretion in patients with type 2 diabetes and impaired glucose tolerance.

The insulinotropic gut hormone glucagon-like peptide (GLP)-1 increases secretory burst mass and the amplitude of pulsatile insulin secretion in healthy volunteers without affecting burst frequency. Effects of GLP-1 on secretory mechanisms in type 2 diabetic patients and subjects with impaired glucose tolerance (IGT) known to have impaired pulsatile release of insulin have not yet been studied. Eight type 2 diabetic patients (64+/-9 years, BMI 28.9+/-7.2 kg/m2, HbA1c 7.7+/-1.3%) and eight subjects with IGT (63+/-10 years, BMI 31.7+/-6.4 kg/m2, HbA1c 5.7+/-0.4) were studied on separate occasions in the fasting state during the continued administration of exogenous GLP-1 (1.2 pmol x kg(-1) x min(-1), started at 10:00 P.M. the evening before) or placebo. For comparison, eight healthy volunteers (62+/-7 years, BMI 27.7+/-4.8 kg/m2, HbA1c 5.4+/-0.5) were studied only with placebo. Blood was sampled continuously over 60 min (roller-pump) in 1-min fractions for the measurement of plasma glucose and insulin. Pulsatile insulin secretion was characterized by deconvolution, autocorrelation, and spectral analysis and by estimating the degree of randomness (approximate entropy). In type 2 diabetic patients, exogenous GLP-1 at approximately 90 pmol/l improved plasma glucose concentrations (6.4+/-2.1 mmol/l vs. placebo 9.8+/-4.1 mmol/l, P = 0.0005) and significantly increased mean insulin burst mass (by 68%, P = 0.007) and amplitude (by 59%, P = 0.006; deconvolution analysis). In IGT subjects, burst mass was increased by 45% (P = 0.019) and amplitude by 38% (P = 0.02). By deconvolution analysis, insulin secretory burst frequency was not affected by GLP-1 in either type 2 diabetic patients (P = 0.15) or IGT subjects (P = 0.76). However, by both autocorrelation and spectral analysis, GLP-1 prolonged the period (lag time) between subsequent maxima of insulin concentrations significantly from approximately 9 to approximately 13 min in both type 2 diabetic patients and IGT subjects. Under placebo conditions, parameters of pulsatile insulin secretion were similar in normal subjects, type 2 diabetic patients, and IGT subjects based on all methodological approaches (P > 0.05). In conclusion, intravenous GLP-1 reduces plasma glucose in type 2 diabetic patients and improves the oscillatory secretion pattern by amplifying insulin secretory burst mass, whereas the oscillatory period determined by autocorrelation and spectral analysis is significantly prolonged. This was not the case for the interpulse interval determined by deconvolution. Together, these results suggest a normalization of the pulsatile pattern of insulin secretion by GLP-1, which supports the future therapeutic use of GLP-1-derived agents.

Aged↗
PubMedDate not supplied

Quantitative measurement of wall shear rate by pulsed Doppler ultrasound.

A 20 MHz pulsed Doppler ultrasound system was used to obtain steady flow velocity profiles in cylindrical tubes. A deconvolution technique was employed to improve the spatial resolution and the corrected results were compared with Poiseuille's theory. Frequency domain windowing was used to suppress noise in the deconvolution procedure, using a method which reduced distortions at the window boundaries. Wall shear rate was calculated from the measured velocity profiles both before and after deconvolution, and compared with theory to quantify the accuracy of the technique. The results illustrate the significant improvement provided by deconvolution, with the average error in measured wall shear rate reducing from 46 to 12% after applying the technique. Some of the discrepancy is in part attributed to the method used to determine wall shear rate.

Biomechanical Phenomena↗
PubMedDate not supplied

Quantitative evaluation of liver function with MRI Using Gd-EOB-DTPA.

OBJECTIVE: Gadolinium ethoxybenzyl diethylenetriaminepentaacetic acid (Gd-EOB-DTPA) is a newly developed MR contrast agent. After intravenous injection, Gd-EOB-DTPA is gradually taken up by the hepatocytes and eventually excreted via the biliary pathway without any change to its chemical structure. Because of these characteristics, it can be used as a tracer for quantitative liver function testing. The purpose of this study is to develop a noninvasive method of quantitation of the hepatic function using Gd-EOB-DTPA through the deconvolution analysis. MATERIALS AND METHODS: Adult New Zealand white rabbits (n = 10, average body weight = 3.5 kg) were used in the present study. Hepatic injury was induced to by the intragastric administration of carbon tetrachloride (CCl4) three times a week for three weeks. Liver enzyme (aspartate aminotransferase, AST; alanine aminotransferase, ALT) levels and the plasma indocyanine green (ICG) retention rate 15 minutes after an intravenous injection of ICG (ICG R15), was checked before and after the three-week administration of CCl4. At the end of experimental period, an observer "blinded" to the treatment given the rabbits performed the histological examination. MRI studies were performed before and after the three-week administration of CCl4 on a 1.5 T scanner using a human extremity coil. After intravenous bolus injection of Gd-EOB-DTPA (0.3 mL of Gd-EOB-DTPA freshly prepared in 2.7 mL of normal saline) through the ear vein, the 250 axial single level dynamic MR images were obtained using a fast low angle shot (FLASH, T /TE = 11/4.2 msec, flip angle = 15, acquisition time 1 second, slice thickness = 5 mm, matrix = 128x128, field of view = 120 mm) sequence with 1.5 sec time intervals. The time-intensity curves were obtained at the abdominal aorta and the liver parenchyma that was devoid of blood vessels. Deconvolution analysis of the aortic (input function) and hepatic parenchymal (output function) time-intensity curves was performed with a modified Fourier transform technique to calculate the hepatic extraction fraction (HEF). The presence and type of hepatic injury were determined by the histopathologic examination and statistical analysis of the changes of the hepatic enzyme levels, the ICG R15 and Gd-EOBDTPA HEF values between the time before and after CCl4 administration with Wicoxon signed rank test. Correlation between the Gd-EOB-DTPA HEF and the change of the ICG R15 were analyzed with Pearson's correlation coefficient. RESULTS: Histopathologic examination showed findings that were compatible with hepatic fibrosis caused by chronic liver injury. The initial blood biochemical studies before the administration of carbon tetrachloride showed that the mean AST and ALT levels were 39.8+/-5.2 IU/L and 59.1+/-11.7 IU/L, respectively. The AST and ALT levels increased to 138.4+/-50.5 IU and 172.0+/-71.6 IU/L, respectively, after the three week administration of CCl4. The ALT and AST levels were significantly increased after the three weeks of CCl4 administration (p = 0.018). The ICG R15 values were 4.47+/-2.08% and 19.43+/-3.98% before and after three-week administration of CCl4, respectively. The ICG R15 values were significantly increased after hepatic injury (p = 0.018). After normalizing the HEF as 100% in each rabbit before CCl4 administration, the deconvoluted curve after CCl4 administration revealed less hepatocyte extraction efficiency with a mean value of 77.7+/-3.6. There was a significant correlation between the HEF and changes of the ICG R15 by the Pearson correlation coefficient assessment (correlation coefficient = -0.965, p = 0.000). CONCLUSION: The Gd-EOB-DTPA HEF could be calculated from deconvolution analysis of aortic and hepatic parenchymal time-intensity curves obtained by dynamic MRI. The Gd-EOB-DTPA HEF was well correlated with changes of the ICG R15, which is the most common parameter used in the quantitative estimation of the hepatic function. The Gd-EOB-DTPA HEF is a direct, noninvasive technique for the quantitative evaluation of liver function. It could be a promising alternative for the determination of noninvasive hepatic function in those patients with liver disease.

Alanine Transaminase↗
PubMed2006

High-resolution ultrasonic imaging using fast two-dimensional homomorphic filtering.

A new method for two-dimensional deconvolution of medical ultrasonic images is presented. The spatial resolution of the deconvolved images is much higher compared to the common images of the fundamental and second harmonic. The deconvolution also results in a more distinct speckle pattern. Unlike the most published deconvolution algorithms for ultrasonic images, the presented technique can be implemented using currently available hardware in real-time imaging, with a rate up to 50 frames per second. This makes it attractive for application in the current ultrasound scanners. The algorithm is based on two-dimensional homomorphic deconvolution with simplified assumptions about the point spread function. Broadband radio frequency image data are deconvolved instead of common fundamental harmonic data. Thus, information of both the first and second harmonics is used. The method was validated on image data recorded from a tissue-mimicking phantom and on clinical image data.

Algorithms↗
PubMed2006

A 'bottom-up' approach for endo-PK/PD analysis.

A 'bottom-up' PK/PD analysis approach employing system analysis principles of convolution/deconvolution and special nonparametric estimation procedures is presented to resolve the complex 'endo-PK/PD' of the endogenous form of recombinant drugs using erythropoietin (EPO) as an example. A novel cellular deconvolution algorithm is presented that facilitates the identification of the functional relationship between the variables involved in EPO's complex PK/PD. Five sheep each underwent two phlebotomies spaced 4-6 weeks apart when their hemoglobin levels were reduced from 12 g/dl to 3-4 g/dl. EPO levels and reticulocyte counts were frequently sampled. The data were analysed using end-constrained cubic splines. The rate of reticulocyte production was determined using the novel deconvolution methodology. The erythroid progenitor cells activation rate by EPO was estimated from the reticulocyte production rate using a lag-time parameter which determines the delay in the reticulocyte appearance in the blood relative to the activation of erythroid progenitors. Hysteresis minimization combined with cellular deconvolution was employed to determine the population PK/PD transduction function relating the progenitor activation rate to EPO concentrations in a nonparametric manner without assuming a specific structure. The proposed approach provides a rational informative starting point for developing parametric PK/PD models to resolve the complex endo-PK/PD of recombinant drugs.

Anemia↗
PubMedDate not supplied

Nature of alpha subunit secretion in men: circadian rhythms, pulsatile release and secretory profiles.

Alpha subunit complements LH as a marker of the activity of the hypothalamic GnRH pulse generator. To characterize episodes of alpha subunit release and to determine if a circadian pattern of alpha subunit secretion is present in man, spontaneous alpha subunit pulsatility was analyzed in six healthy young men by blood sampling every 5 min for 24 h. The resulting alpha subunit concentration time series were analyzed by two statistically based independent peak detection methods, and subjected to Fourier transformation to assess underlying circadian rhythms. Cross-correlation analyses and multiple parameter deconvolution were used to estimate the concordance of spontaneous and exogenous GnRH-stimulated LH and alpha subunit secretion. These analyses revealed that two independent discrete peak detection algorithms yielded similar estimates of spontaneous alpha subunit pulse frequency, namely, 21 +/- 1.1 (Cluster) and 21 +/- 1.5 (Detect) alpha subunit peaks/24 h. Sampling intensity markedly influenced the estimate of endogenous alpha subunit pulse frequency, inasmuch as estimates from 5-min sampling were significantly greater than those of 10-min or 20-min sampling. Fourier transformation unmasked a significant circadian alpha subunit rhythm in all six men, with maximal concentrations at 0836 h and an average amplitude of 28% of the 24-hr mean hormone concentration. Cross-correlation analysis of spontaneous glycoprotein release revealed that serum LH and alpha subunit concentrations were highly cross-correlated when considered simultaneously, but not at various lags. Finally, deconvolution analysis of exogenous GnRH-stimulated glycoprotein release disclosed distinct half-times of alpha subunit and LH clearance with virtually simultaneous underlying secretory bursts. These data indicate that human alpha subunit is secreted in both a circadian and a discrete pulsatile fashion at a pulse frequency that is significantly underestimated at conventional sampling rates. The approximately hourly alpha subunit interpulse interval (68 +/- 4.6 min) is similar to that reported earlier for LH in peripheral blood and for testosterone in gonadal vein blood in healthy men. Moreover, cross-correlation analysis of endogenous GnRH-driven alpha subunit and deconvolution analysis of exogenous GnRH-stimulated alpha subunit and LH secretion suggest that these glycoproteins are secreted virtually simultaneously, but have significantly different endogenous clearance properties. The remarkably similar in vivo pulse frequencies for alpha subunit, LH, and testosterone in man suggest that the release of these three hormones is coordinately regulated.

Adult↗
PubMed2004

Colocalization analysis yields superior results after image restoration.

Colocalization analysis is a powerful tool for the demonstration of spatial and temporal overlap in the distribution patterns of fluorescent probes. In unprocessed images, background affects image quality by impairing resolution and obscuring image detail in the low-intensity range. Because confocal images suffer from background levels up to 30% maximum intensity, colocalization analysis, which is a typical segmentation process, is limited to high-intensity signal. In addition, noise-induced, false-positive events ("dust") may skew the results. Therefore, suppression of background is crucial for this type of image analysis. Analysis of synthetic and biological objects demonstrates that median filtering is able to eliminate noise-induced colocalization events successfully. Its disadvantages include the occasional generation of false-positive and false-negative results as well as the inherent impairment of resolution. In contrast, image restoration by deconvolution suppresses background to very low levels (<10% maximum intensity), which makes additional objects in the low-intensity but high-frequency range available for analysis. The improved resolution makes this technique extremely suitable for examination of objects of near resolution size as demonstrated by correlation coefficients. Deconvolution is, however, sensitive to overestimation of the background level. Conclusions for practical application are: (1) In raw images, colocalization analysis is limited to the intensity range above the background level. This means the higher the RS/N the better. Unfortunately, images of most biological specimens have a low RS/N. (2) Filtering improves the result substantially. The reduction of background levels and the concomitant increase of the RS/N are generated at the expense of resolution. This is a quick and simple method in cases where resolution is not a major concern. (3) If colocalization in the low-intensity range and/or maximum resolution play a role, deconvolution should be used.

Animals↗