Quantitative method for in vitro matrigel invasiveness measurement through image analysis software

The determination of cell invasion by matrigel assay is usually evaluated by counting cells able to pass through a porous membrane and attach themselves to the other side, or by an indirect quantification of eluted specific cell staining dye by means of optical density measurement. This paper describes a quantitative analytical imaging approach for determining the invasiveness of tumor cells using a simple method, based on images processing with the public domain software, ImageJ. Images obtained by direct capture are split into the red channel, and the generated image is used to measure the area that cells cover in the picture. To overcome the several disadvantages that classical cell invasion determinations present, we propose this method because it generates more accurate and sensitive determinations, and it could be a reasonable option for improving the quality of the results. The cost-effective alternative method proposed is based on this simple and robust software that is worldwide affordable.     

Direct quantification of in vitro cell growth through image analysis

Summary Objective, accurate, non-intrusive measurement of in vitro cell growth was realized through microcomputerized video image analysis. Recently-released video and digitizing hardware and software were incorporated into an analytical system which accurately quantified visual differences between cultures on a cell number or fresh mass basis. Sequential measurements during culture incubation further detected and quantified subtle changes in colony area and density resulting from growth. Each measurement was acquired rapidly, without encroaching on the in vitro environment, so cell growth was undisturbed. Custom software routines coordinated the quantification of this detailed record into precise cumulative growth curves.     

Quantitation of in vitro ciliated cell growth through image analysis

Summary Ciliated cell cultures can be produced in outgrowths from explants of human respiratory epithelium. An image analysis technique was develope to quantify the percentage of active ciliated cells present in these cultures. The subtraction 2 by 2 of five successive video images of the cultures, followed by the addition of the resulting images, allowed the determinaton of the culture surface covered by ciliated cells. The percentage of this surface varied according to the regions of the explant (27.7% in the outgrowth near the explant and 4.1% at the periphery of the outgrowth). High variations were observed within the same region of an outgrowth, as well as from one outgrowth to another. However, maximal differentiation was observed after 4 d of culture. The quantitation techniques described in the present work might be useful for studying in vitro the respiratory epithelial injury and the subsequent repair processes. This work was supported by CEB-INSERM and SYNTHELABO-INSERM grants.     

Finding an image in a haystack: the case for public image repositories

Image repositories are growing. Their potential impact is huge; they require support and proper funding.     

plusTipTracker: Quantitative image analysis software for the measurement of microtubule dynamics

Here we introduce plusTipTracker, a Matlab-based open source software package that combines automated tracking, data analysis, and visualization tools for movies of fluorescently-labeled microtubule (MT) plus end binding proteins (+TIPs). Although +TIPs mark only phases of MT growth, the plusTipTracker software allows inference of additional MT dynamics, including phases of pause and shrinkage, by linking collinear, sequential growth tracks. The algorithm underlying the reconstruction of full MT trajectories relies on the spatially and temporally global tracking framework described in Jaqaman et al. (2008). Post-processing of track populations yields a wealth of quantitative phenotypic information about MT network architecture that can be explored using several visualization modalities and bioinformatics tools included in plusTipTracker. Graphical user interfaces enable novice Matlab users to track thousands of MTs in minutes. In this paper, we describe the algorithms used by plusTipTracker and show how the package can be used to study regional differences in the relative proportion of MT subpopulations within a single cell. The strategy of grouping +TIP growth tracks for the analysis of MT dynamics has been introduced before (Matov et al., 2010). The numerical methods and analytical functionality incorporated in plusTipTracker substantially advance this previous work in terms of flexibility and robustness. To illustrate the enhanced performance of the new software we thus compare computer-assembled +TIP-marked trajectories to manually-traced MT trajectories from the same movie used in Matov et al. (2010).     

The use of automated image analysis for rapid measurement of the in vitro attachment of Candida albicans to transparent acrylic

A rapid method for measuring the in vitro attachment of Candida albicans to the surface of transparent acrylic is described. The method involves use of the 'Magiscan' automated image analysis system which measures attachment in terms of percentage area coverage. These measurements correlate highly significantly ( P < 0·001) with the number of adherent yeast cells.     

Morphometric requirements for image analysis and the IBAS interactive automatic image analysis system

The morphometric demands made on an image analysis system are discussed, as is the IBAS interactive automatic system, which meets those requirements. The modular design of the IBAS image analysis system, with its tailor-made processors for image processing, system control and pattern recognition, gives speed and flexibility. The IMAGE image analysis language guarantees user friendliness, and, last but not least, the enormous amount of software offers accurate, reproducible measurements and dedicated evaluation programs.     

In vitro analysis of the complete CIDE domain interactions of the Drep system in fly

CIDE domain containing proteins are involved in apoptosis and lipid metabolism, and four CIDE containing proteins, Drep1, Drep2, Drep3, and Drep4, have been identified in fly. In this study, we found that Drep3 interacts with Drep4 via the CIDE domain specifically, which completes the interaction map of Drep system in fly, cyclic interactions: Drep1–Drep2–Drep3–Drep4–Drep1. In addition, we analyzed the dynamic stoichiometry changes of Drep proteins upon binding to their binding partners. Our current studies will help us to understand Drep system in fly as well as CIDE domain for protein–protein interactions.     

Robust framework for PET image reconstruction incorporating system and measurement uncertainties

In Positron Emission Tomography (PET), an optimal estimate of the radioactivity concentration is obtained from the measured emission data under certain criteria. So far, all the well-known statistical reconstruction algorithms require exactly known system probability matrix a priori, and the quality of such system model largely determines the quality of the reconstructed images. In this paper, we propose an algorithm for PET image reconstruction for the real world case where the PET system model is subject to uncertainties. The method counts PET reconstruction as a regularization problem and the image estimation is achieved by means of an uncertainty weighted least squares framework. The performance of our work is evaluated with the Shepp-Logan simulated and real phantom data, which demonstrates significant improvements in image quality over the least squares reconstruction efforts.     

Performance of a TV image analysis system as a microdensitometer

The performance of a TV image analysis system combined with an automated microscope (the Leitz TAS plus) as a microdensitometer and morphometric device was investigated. There was a strict linear correlation (r greater than .99) between the physically defined optical transmission values and the resulting electronic signals from the plumbicon TV camera for the whole area displayed on the monitor. The shading of the optoelectronic system had a coefficient of variation (CV) of 1.42% for measurements in the center of the displayed area, but a CV of 3.55% for measurements over the whole monitor area. Densitometric measurements remained stable 15 minutes after putting the microscope lamp into operation (T less than 0.075%, remeasuring every two minutes). The geometric distortion, measured as different ferret diameters of ideally round latex particles, ranged from +/- 0.5% to +/- 1.0% deviation over the entire displayed area. These results indicate that densitometric and morphometric measurements with this equipment are sufficiently precise and reproducible when performed in the center of the area displayed on the monitor.     

In vitro measurement of pollen tube growth inhibition

A method for estimating inhibition of pollen tube growth was developed. Pollen is placed in straight lines on an agar surface where it responds uniformly and predictably to aqueous solutions of germination-inhibiting substances located in wells at the ends of the lines. A scale of ratings, roughly corresponding to serial, doubled concentrations of inhibiting substances, was devised. Water-soluble organic solvents are relatively noninhibitory, salts are variable, and metabolic inhibitors have strong inhibitory effects. Pollens differ in their susceptibility to inhibition and in their response to particular substances.     

Iron formulation affects in vitro storage of hops: An image analysis

Summary In vitro-stored plant germplasm is usually evaluated by visual analysis of the plant based on subjective characters. To reduce the variability in these evaluations, we developed a digital-image evaluation system for in vitro-stored plantlets. This study compares the standard visual evaluation system with a digital analysis system to determine if digital analysis can effectively quantify the health of diverse Humulus germplasm. Eight cultivars of Humulus lupulus L. were stored on standard Murashige and Skoog (MS) medium with iron alone (EDTA chelated) and on MS iron with 100 or 200 mgl^&#x2212;1 sequestrene 138 iron (EDDHA chelated). Digital images of the upper two nodes of each plantlet were evaluated for red, green, blue, green/red ratio, and modified normalized difference vegetation index (MNDVI=R-G/R+G). Evaluation of each plantlet for MNDVI values showed consistent significant differences for all treatments only at the upper node. Significant differences for visual and the MNDVI values among the three iron treatments were observed at the upper node of most of the eight hop cultivars. Regression analysis of the upper node MNDVI values vs. whole-plant visual ratings showed positive correlations for most cultivars. Effects of iron treatments on storage duration were also analyzed for both visual and digital systems. There were significant differences among MNDVI values for plantlets stored on medium with standard MS iron alone (EDTA chelated) and with the addition of sequestrene 138 iron. In general, the MNDVI value of the upper node correlated well with visual ratings and could be used to determine the health of in vitro stored hops.     

A Computer-based image analysis system for biocide screening

A computer-based image analysis system that measures the behavioral response of the marine diatom Amphora coffeaeformis to potentially toxic challenges is described. At any one time the system is capable of measuring in a population of about 80 cells, changes in the number motile, their speed, direction, and if they are turning, their angular velocity and acceleration. Chemical compounds that interfere with the generation of energy, protein or glycoprotein synthesis, calcium homeostasis and cytoskeletal activity, change the motile behavior of the cell. A pre-requisite of motility is that the cells be attached to a substratum. It is proposed that the system, which requires only a 1-2 minute data collection period per replicate, could be used to screen for potential antifouling activity in chemical agents, and it is speculated that it could be used also for measuring the physicochemical interaction between attached motile organisms and their substratum, as well as in environment toxicology measurements.     

An automatic image analysis system for RLGS films

The RLGS (Restriction Landmark Genome Scanning) method was originally developed as a powerful method for enabling viewing of thousands of restriction landmarks. It offers a tool for obtaining information about genetic loci, with a single RLGS profile displaying approximately 2000 restriction landmarks as spots. One of the most useful applications is RLGS spot mapping, which allows the efficient, low-cost construction of the genetic map of any organism. However, analyses of the profiles depend mainly on human visual observation and are tedious and laborious. Although several commercially available image analyzing systems for profile comparison have been examined, they cannot be used for the RLGS spot mapping system owing to the background characteristics of the RLGS profiles, unsatisfactory rates of correspondence, and inefficient correction of informative genetic data. We therefore developed a novel automatic image analysis system for RLGS spot mapping, using an original algorithm based on the binary image transferred from the original RLGS profile. This system was employed for identifying non-polymorphic and parental strain-specific polymorphic spots of the F₁ mouse profile and yielded efficient initial screening of RLGS profiles. Received: 5 December 1997 / Accepted: 6 April 1998     

Photosynthetic competence of plantlets grown in vitro. An automated system for measurement of photosynthesis in vitro

An aseptic gas exchange and hydroponic system (AGEHS) has been developed in an attempt for characterization of physiological requirements for photoautotrophic growth in vitro and alleviation of the needs for ex vitro acclimatization. The AGEHS monitors and controls several parameters relevant to plant growth. Shootlets of Chrysanthemum × morifolium Ramat. cv. Envy were treated with flow of air or CO₂-enriched air under controlled relative humidity, elevated photosynthetic photon flux density (PPFD) and hydroponic irrigation. After 15 days of treatment, plantlets gained more than 3 times as much dry weight as those from a conventional culture tube treatment. This study shows that it is possible to favour photoautotrophic growth when elevated PPFD, enhanced air-exchange and hydroponic medium flow are provided concurrently. This enhancement is achievable through careful increments of light quanta, balanced with increments of humidified air flow and/or CO₂ content in air which seem to be necessary to avoid potential photoinhibition and premature water exhaustion from gelled media.     

Local and global cycles in an acarine predator-prey system: a frequency domain analysis

Techniques of time series analysis are applied to the dynamics of the phytophagous miteTetranychus urticae Koch and its predatorTyphlodromus occidentalis Nesbitt in six experimental mini-orchards, sampled weekly during two years (van de Klashorst et al., 1992). Autocovariance and crosscovariance functions characterize local and global behaviour in the time domain. Spectral density function and cross amplitude spectrum provide important information of the system's behaviour in the frequency domain. Because of a strikingly different behaviour, the total record was cut into two distinct periods. During the first period, all local systems oscillate with a frequency of four cycles per year and all in phase, resulting in a strongly periodical global behaviour at a high mean level. Since the mean local power spectra coincide with the global power spectrum over a wide range of frequencies, it is concluded that the total system of six orchards is homogeneous. During the second period, all local power spectra are mutually different, more or less smoothed with no apparent peak pointing to a periodical component. The resulting global power spectrum is almost flat at a low mean level. The system is heterogeneous with a quasi-stable global behaviour.From the cross-amplitude spectrum it became clear that the dynamics of the two species within the orchards remained strongly coupled over a wide range of frequencies.Since the experimental circumstances had not been intentionally changed, the origin of the drastic change in behaviour could not be identified. If high rates of dispersal have been the synchronizer during the first period, it is not clear why asynchrony suddenly occured in the second period.A comparison of the overall record with the results of Nachman's (1987) stochastic simulation model suggests that the change could possibly be of stochastic origin.