Reconfigurable modular automation systems for automotive power-train manufacture

This paper describes research towards the realization of reconfigurable modular automated machines and the associated engineering methods and tools necessary to support their lifecycle needs. UK-based research, in collaboration with the Ford Motor Company and several machine builders, has resulted in the development of full-scale prototype reconfigurable modular automation systems for both engine assembly and machining applications. The implementation of an assembly system is featured in this paper. An engineering environment and associated reconfigurable component-based control system architecture have been created aimed at supporting the lifecycle needs of a new generation of agile automated systems, i.e., providing reconfigurable, easily scalable automated machinery. This approach has the potential to fit within a wider collaborative automation strategy where manufacturing systems are implemented as a conglomerate of distributed, autonomous, and reusable units.     

"6 markers/5 colors" extended white blood cell differential by flow cytometry.

Electronic white blood cell (WBC) differential by standard cytology (hematology analyzer and visual inspection of blood smears) is limited to five types and identification of abnormal cells is only qualitative, often problematic, poorly reproducible, and labour costing. We present our results on WBC differential by flow cytometry (FCM) with a 6 markers, 5 colors CD36-FITC/CD2-PE+CRTH2-PE/CD19-ECD/CD16-Cy5/CD45-Cy7 combination, on 379 subjects, with detection of 12 different circulating cell types, among them 11 were quantified. Detection of quantitative abnormalities of whole leucocytes, neutrophils, eosinophils, basophils, monocytes, or lymphocytes was comparable by FCM and by standard cytology in terms of sensitivity and specificity. FCM was better than standard cytology in detection and quantification of circulating blast cells or immature granulocytes, with a first lineage orientation in the former case. All cases of lymphocytosis, with lineage assignment, were detected by FCM. FCM identified a group of patients with excess of CD16pos monocytes as those having an inflammatory syndrome. WBC differential by FCM is at least as reliable as by standard cytology. FCM superiority consists in identification and systematic quantification of parameters that cannot be assessed by standard cytology such as lineage orientation of blast cells or lymphocytes, and expression of markers of interest such as CD16 on inflammatory monocytes.     

Functional proteometrics for cell migration.

BACKGROUND: Advances in living cellular fluorescence biosensors and computerized microscopy enable a vision of fully automated high-resolution measurements of the detailed intracellular molecular dynamics directly linked to cellular behaviors. Given the heterogeneity of cell populations, a statistically relevant study of molecular-cellular dynamics is a key motivation for improved automation. METHODS: We explored automating computerized, microscope-based data extraction and analyses that monitor cell locomotion, rates of mitoses, and spatiotemporal activities of intracellular proteins via ratiometric fluorescent biosensors in mouse fibroblasts. Novel image processing methods included K-means clustering segmentation preprocessing followed by modified discrete, normalized cross-correlational alignment of two-color images; ratiometric processing for fluorescence resonance energy transfer (FRET) measurements; and intracellular spatial distribution measurements of RhoA GTPase activity. RESULTS: The interdivision time was 19.4 h (mean) +/- 6.0 h (SD) (n = 7) for the GFP-histone cells in the two-by-two field that was scanned for 72 h. After registration and ratioing of the cells with the RhoA biosensor, increases in both cell protrusion and retraction were coincident with to increases in RhoA activity. CONCLUSIONS: These advances lay the foundation for extracting and correlating measurements characterizing the functional relationships of spatial localization and protein activation with features of cell migration such as velocity, polarization, protrusion, retraction, and mitosis.     

Automation of the analysis of cell images

This paper reviews the advances made in automated cytology during the past three decades, with particular attention to the development of white blood cell image differentiation equipment. The currently available commercial systems for white blood cell recognition are described in detail.     

Automation of immunohematologic testing activities at French blood transfusion centers

In May 1982, a questionnaire was sent to all of the 170 French Blood Transfusion Services (BTS), on behalf of the French Society of Blood Transfusion. The purpose was to determine the types of automated equipment used for immunohematological controls, the way in which they are used and the result of automation and computerization in daily laboratory operations. We received 135 replies (80%). A generalized conclusion can be drawn from the collected information. 50% of the respondents are neither automated nor computerized. 30% are both automated and computerized. 10% are automated but not computerized and 8% are not automated but are computerized. In the field of automated serology there is an increased tendency to complete the ABO/Rh testing by Cc D Ee and Kell phenotyping. The use of computers allows the current test determination to be compared with previous donation data. However, no fully automated equipment, which can conduct antibody screening, exists, cost effectively, in small or average BTS. In France, there has been a significant increase in automation between 1970 and 1980 but only the most important BTS have carried out automation at the same time as computerization. The smaller BTS have usually become automated without becoming computerized. In 1978, Codabar was first used. This has been one of the principal advances of the last 10 years, allowing all the users of automation to start moving towards complete computerization. This advance was assisted by the use of prepackaged software. This questionnaire also determined that the current emphasis is now to computerize administrative and management activities before laboratory activities. This survey has been conducted during a turning point of the automation of French BTS. It shows that they are, on the whole, satisfied with their automation. As far as the safety and the efficiency of the service are concerned, it is only fair to consider that the main purposes of the automation have been achieved. But in terms of cost, and serological accuracy for antibody screening, a new generation of automated equipment should appear to satisfy the users in the nineties.     

Performance of a cost‐effective and automated blood counting system for resource‐limited settings operated by trained and untrained users

Current flow‐based blood counting devices require expensive and centralized medical infrastructure and are not appropriate for field use. In this article we report a streamlined, easy‐to‐use method to count red blood cells (RBC), white blood cells (WBC), platelets (PLT) and 3‐part WBC differential through a cost‐effective and automated image‐based blood counting system. The approach consists of using a compact, custom‐built microscope with large field‐of‐view to record bright‐field and fluorescence images of samples that are diluted with a single, stable reagent mixture and counted using automatic algorithms. Sample collection utilizes volume‐controlled capillary tubes, which are then dropped into a premixed, shelf‐stable solution to stain and dilute in a single step. Sample measurement and analysis are fully automated, requiring no input from the user. Cost of the system is minimized through the use of custom‐designed motorized components. We compare the performance of our system, as operated by trained and untrained users, to the clinical gold standard on 120 adult blood samples, demonstrating agreement within Clinical Laboratory Improvement Amendments guidelines, with no statistical difference in performance among different operator groups. The system's cost‐effectiveness, automation and performance indicate that it can be successfully translated for use in low‐resource settings where central hematology laboratories are not accessible.      

Automated cervical cytology screening: performance requirements and instrument evaluation.

Continuing progress towards automation of cervical cancer screening requires that criteria for clinical acceptability of automated systems be defined, and that methods be devised for effectively evaluating new technology. The potential roles of automation in cervical cancer detection, performance requirements, instrument evaluation and useful contributions from tuberculosis screening, automated white blood cell differential counting, signal detection theory and decision theory are discussed.     

O-10THE ACCURACY OF CERVICAL CYTOLOGY: A COMPARISON BETWEEN THE THINPREP IMAGING SYSTEM AND CONVENTIONAL METHODS

Douglass Hanly Moir is a large Australian laboratory which has recently introduced the ThinPrep Imaging System (TPI) for reading ThinPrep slides, which is still performed using a split-sample technique. The Imager is a computerized system which identifies 22 fields for the cytologist to review using automated light microscopy. We compared the accuracy of TPI and conventional cytology (CC) during normal laboratory operation. The ThinPrep sample was prepared after taking a conventional Pap smear. TPI and CC reading was done without knowledge of the result of the other reading. The final cytology report issued to the referring doctor reflected the more severe of these two results. Histology results for all cases in which TPI and CC cytology results showed more than minimal disagreement were sought from the NSW Pap Test Register. Of 55 164 split sample pairs, 3.1% of CC of slides and 1.8% of TPI slides were unsatisfactory. There were 1758 women for whom there was more than minimal discrepancy between TPI and CC cytology results. TPI gave the more severe result in 1193 of the 1758 cases. In cases where only one of each pair of discrepant cytology results was CIN1 or higher grade, TPI detected 133 cases of high-grade histology among 380 biopsies (35%), whereas CC detected 62 cases among 210 biopsies (29.5%). A repeat analysis based on reading of histology by one pathologist blinded to initial Pap smear result showed a similar result. Reading times were measured over 5 months for both TPI and CC for twenty cytologists who read both types of smears. On average, they read 13.3 TPI slides per hour and 6.1 CC slides per hour. This study provides evidence that cervical cytology read using the TPI detects more histological high-grade disease than does CC. Further evidence shows that reading times are significantly reduced for cytologists using the TPI.     

FDA regulation of computerized cytology devices

When talking about computerized cytology devices, a "different" aspect of quality assurance must be addressed. Any medical device intended for in vitro diagnostic use in the United States must be cleared or approved by the Food and Drug Administration (FDA): the May 28, 1976, Medical Device Amendments to the Federal Food, Drug and Cosmetic Act granted authority to the FDA to regulate medical devices. The FDA regulatory process as it relates to computerized cytology devices is discussed. This includes an explanation of the differences between the two types of documents used to clear a medical device: (1) premarket notification [510(k)] and (2) premarket approval (PMA) application. Devices intended for "research use only" are also discussed. A computerized cytology device of current interest, the "automated Pap smear reader," is used as an example to further discuss performance and software considerations.     

MalariaCount: an image analysis-based program for the accurate determination of parasitemia

Malaria is a serious global health problem and rapid, precise determination of parasitemia is necessary for malaria research and in clinical settings. Manual counting by light microscopy is the most widely used technique for parasitemia determination but it is a time-consuming and laborious process. The aim of our study was to develop an automated image analysis-based system for the rapid and accurate determination of parasitemia. We have developed, for the first time, a software, MalariaCount, that automatically generates parasitemias from images of Giemsa-stained blood smears. The potential application and robustness of MalariaCount was tested in normal and drug-treated in vitro cultures of Plasmodium falciparum. The results showed a tight correlation between MalariaCount and manual count parasitemia values. These findings suggest that MalariaCount can potentially be used as a tool to provide rapid and accurate determination of parasitemia in research laboratories where frequent, large-scale, efficient determination of parasitemia is required.     

Multi-resolution correlative focused ion beam scanning electron microscopy: Applications to cell biology

Efficient correlative imaging of small targets within large fields is a central problem in cell biology. Here, we demonstrate a series of technical advances in focused ion beam scanning electron microscopy (FIB–SEM) to address this issue. We report increases in the speed, robustness and automation of the process, and achieve consistent z slice thickness of ∼3 nm. We introduce “keyframe imaging” as a new approach to simultaneously image large fields of view and obtain high-resolution 3D images of targeted sub-volumes. We demonstrate application of these advances to image post-fusion cytoplasmic intermediates of the HIV core. Using fluorescently labeled cell membranes, proteins and HIV cores, we first produce a “target map” of an HIV infected cell by fluorescence microscopy. We then generate a correlated 3D EM volume of the entire cell as well as high-resolution 3D images of individual HIV cores, achieving correlative imaging across a volume scale of 10⁹ in a single automated experimental run.     

Automatic leukocyte classification using cytochemically stained smears.

A leukocyte classification algorithm suitable for automated differential counting has been developed for blood smears stained with a new three-component cytochemical stain which has relatively narrow absorption bands centered at 460, 540 and 640 nm, respectively. The classification procedure is the result of a pattern recognition experiment using a sample of 223 leukocytes distributed evenly over the five normal cell types. The basic data for each cell were three digital microscopic images obtained with narrow band illumination at the above central wavelengths using a TV-digitizer system interfaced to a PDP-15 computer. The classification algorithm involves a sequential decision procedure utilizing five pattern features computed from the intensity histograms of the green and blue digital images. Thus the number of arithmetic operations and the number of computer memory words necessary to perform the classification into one of the five normal white blood cell types are both proportional to n where n is the number of gray levels into which the intensity scale is divided. In this experiment, n equals 256. Comparison of our results with work of others on smears prepared with Romanowski-type stains indicates that such narrow-band, spectrally well separated cytochemical multiple stains can permit the use of algorithms which are approximately ten times faster.     

A Quick and Standardized Giemsa Stain for Wet-Fixed Cytological Material

The Giemsa stain is one of the most widely used staining techniques in cytology, especially in hematology. A standardized Romanowsky-Giemsa staining procedure using pure cationic azure B (C.I. 52010) and anionic eosin (C.I. 45380) has been described by Wittekind et al (1982). A revised standard Giemsa staining procedure was recently published (Wittekind and Kretschmer 1987). Usually the Romanowsky-Giemsa stain is applied to air dried and methanol fixed cytological material, e.g. blood smears and bone marrow films (ICSH 1984).     

Medical applications of image analysis with the Magiscan 2

The previous generation of image analysis machines were capable of processing and analyzing binary, i.e., black and white images, and making measurements and decisions thereon. The Magiscan 2, one of the new generation computers, is capable of analyzing gray-level images in a variety of sophisticated ways. Its uses in the medical application of image analysis are presented, as are the techniques used to analyze images in general. The two principal current medical uses are the automatic karyotyping of chromosomes and the automatic screening of cervical smears. Other applications discussed include three-dimensional reconstruction of structures from two-dimensional sections and the possibility of developing expert systems for medical diagnostics.     

Fluorescent studies directed towards the location of abnormal epithelial cells in cervical smears

Cervical screening is concerned with the search for abnormal epithelial cells in smears prepared from scrapings from the uterine cervix. It is a highly skilled labour intensive operation and automated methods of detecting dyskariotic cells in cervical smears would be helpful. We report a fluorescence method of detecting abnormal cervical cells in smears and biopsies using a probe for guanidinobenzoatase. This approach has the potential for automation.     

Hematologic values of New Zealand white rabbits determined by automated flow cytometry.

Hematologic values of peripheral blood from normal adult New Zealand White rabbits were determined by five different automated flow cytometers in use in a routine clinical hematology laboratory: Technicon H1, Coulter Counter CC540, Coulter Counter VCS, Sysmex NE8000 and Sysmex R1000. The software designed for human blood analysis was used in all instances without adaptation. The total numbers of white blood cells, red blood cells, reticulocytes and platelets were measured with high precision and accuracy. Except for hemoglobin content, concordance was excellent for all measured and calculated values among the different automated flow cytometers. Determining the white blood cell differential count was more complex. Eosinophils and lymphocytes were quantified reliably by all the automated flow cytometers used. However, the results were rejected by Technicon H1 and Sysmex NE8000 in 50% of the cases. Rabbit basophils were recognized with accuracy by Technicon H1 only. The proportion of polymorphonuclear versus mononuclear white cells was identical when measured with Technicon H1 and Coulter Counter VCS. These results show that the new generation of automated flow cytometers designed for human blood can be used with some limitations for animal studies. They allow the standardization of normal values and comparison of results among or between laboratories. They also introduce new parameters, the value of which is as yet undefined.     

华南虎血常规两种测定方法的比较应用

通过全自动血细胞分析仪与人工镜检法分别对华南虎(Panthera tigris amoyensis)血常规进行分析,并对获得结果做比较分析,探究建立适合华南虎血细胞分析的方法。收集临床华南虎血液标本40份,分别使用全自动血液分析仪和人工显微镜检法对红细胞、白细胞进行计数检测,并对白细胞进行分类分析。如人工计数和仪器计数获得的数据均符合正态分布,则应用配对样本T检验比较差异性,否则应用非参数检验(两个相关样本:Wilcoxon带符号秩检验)。相关性采用Spearman分析,以P <0.05为差异有统计学意义。结果表明,通过两种方法获得的红细胞计数、白细胞计数、中性粒细胞比率、嗜酸性粒细胞比率结果差异均不显著(P> 0.05);淋巴细胞比率、单核细胞比率和嗜碱性粒细胞比率,两种方法检测结果具有显著性差异(P <0.05)。红细胞计数(r=0.915)、白细胞计数(r=0.832)、淋巴细胞比率(r=0.832)、中性粒细胞比率(r=0.481)应用两种方法获得的结果相关性均较好。单核细胞比率(r=0.283)、嗜酸性粒细胞比率(r=0.309)、嗜碱性粒细胞比率(r=0....     

Fractal characteristics of May-Grünwald-Giemsa stained chromatin are independent prognostic factors for survival in multiple myeloma

Background

The use of computerized image analysis for the study of nuclear texture features has provided important prognostic information for several neoplasias. Recently fractal characteristics of the chromatin structure in routinely stained smears have shown to be independent prognostic factors in acute leukemia. In the present study we investigated the influence of the fractal dimension (FD) of chromatin on survival of patients with multiple myeloma.

Methodology

We analyzed 67 newly diagnosed patients from our Institution treated in the Brazilian Multiple Myeloma Study Group. Diagnostic work-up consisted of peripheral blood counts, bone marrow cytology, bone radiograms, serum biochemistry and cytogenetics. The International Staging System (ISS) was used. In every patient, at least 40 digital nuclear images from diagnostic May-Grünwald-Giemsa stained bone marrow smears were acquired and transformed into pseudo-3D images. FD was determined by the Minkowski-Bouligand method extended to three dimensions. Goodness-of-fit of FD was estimated by the R² values in the log-log plots. The influence of diagnostic features on overall survival was analyzed in Cox regressions. Patients that underwent autologous bone marrow transplantation were censored at the day of transplantation.

Principal Findings

Median age was 56 years. According to ISS, 14% of the patients were stage I, 39% were stage II and 47% were stage III. Additional features of a bad prognosis were observed in 46% of the cases. When stratifying for ISS, both FD and its goodness-of-fit were significant prognostic factors in univariate analyses. Patients with higher FD values or lower goodness-of-fit showed a worse outcome. In the multivariate Cox-regression, FD, R², and ISS stage entered the final model, which showed to be stable in a bootstrap resampling study.

Conclusions

Fractal characteristics of the chromatin texture in routine cytological preparations revealed relevant prognostic information in patients with multiple myeloma.     

Preparation of monolayer smears from paraffin-embedded tissue for image cytometry

A method is described for the preparation of monolayer smears from paraffin-embedded tissue suitable for automated image analysis and DNA measurements. The proposed technique uses enzyme treatment and syringing for cell dispersal. Slide preparation is performed by centrifugal cytology. After Feulgen staining the quality of the monolayer smears is sufficiently high to enable visual morphologic evaluation. Automated DNA measurements using the Leyden television analysis system (LEYTAS) show coefficients of variation (CV) of 4.5% for the diploid cell population of the suspended tissue. This is approximately the same as the CV in fresh material from the same tumor. Formalin fixed trout red blood cells are used as reference cells. By applying image cytometry to paraffin-embedded tissue this method allows retrospective studies of, for instance, the significance of DNA content with regard to the behavior of a tumor.