Quantitative Digital Analysis Research Articles

Clinical applications of the T-Scan quantitative digital occlusal analysis technology: a systematic review.

Occlusion is associated with all disciplines of dentistry and plays a major role in the longevity of both implant- and tooth-borne restorations. Achieving occlusal harmony ensures balance is established between the dental and myofascial structures, which can be measurably established to high numerical tolerances with the T-Scan digital occlusal analysis system. To describe and evaluate the known and proven applications of T-Scan digital occlusal analysis in various dental practice disciplines through a systematic review of the literature. An electronic, English-language PubMed/MEDLINE and Cochrane Central Registry of Controlled Trials database search using the keywords "T-Scan," "TMD," "Occlusion," "Implant Protected Occlusion," and "Orthodontics" was conducted without any date restrictions. The related journal findings were hand searched to determine studies that met the eligibility criteria for inclusion in the present systematic review. The PubMed/MEDLINE search identified 423 articles. After removing duplicates, the titles and abstracts of the remaining 421 studies were screened. 274 ineligible articles were excluded, leaving 147 articles. Of those, 33 articles were not in English, 27 full-text articles were not available, 4 were comments and letters to editors, 1 was a review, and 2 described techniques. A total of 86 articles met the eligibility criteria for inclusion. Much scientific evidence supports the use of T-Scan, as it measures relative occlusal contact forces and the time sequence durations of occlusal contacts objectively, accurately, and repeatedly for improved treatment outcomes. The system's hardware, sensor, and software evolution from T-Scan I to today's T-Scan 10 Novus system has overcome early sensor and system drawbacks to improve the clinical performance of T-Scan in many disciplines of dental medicine.

Abstract 3652: Spatial transcriptomic study of the tumor microenvironment in HNSCC

Abstract Background: The complex and dynamic nature of the tumor-immune microenvironment (TME) presents challenges for identification of robust and predictive biomarkers in immuno-oncology (IO). Standard multiplex immunohistochemistry (mIHC) or multiplex immuno-fluorescent assays (mIF) allow for the phenotyping on the cellular level but are limited by the number of markers available and the detection systems. Gene expression or next generation sequencing (NGS) platforms provide high-plex bulk data sets about the TME, but the spatial context of the tumor and immune cell interactions is lost. The NanoString GeoMx® Digital Spatial Profiler (DSP) combines spatial context, utilizing fluorescent morphological markers, with molecular profiling capabilities. This combination allows for the quantitative analysis of high-plex analyte abundance which can be traced back to a region of interest within a sample, providing biological inference in the region of interest. Methods: Formalin-fixed paraffin-embedded (FFPE) specimens from HNSCC patients were cut in 5µm sections for all analyses. Whole section tissue analysis was previously performed utilizing the NanoString nCounter® PanCancer IO 360™ Panel. Tumor inflammation signatures (TIS), an 18-gene signature measuring the adaptive immune response within tumors, was generated for each specimen and categorized each as immune-hot or immune-cold based on predefined parameters. Four specimens were selected based on their TIS score and two sets of cut sections were processed by either Labcorp lab in Morrisville, NC or Nanostring lab in Seattle, WA for precision evaluation, using the GeoMx Human Whole Transcriptome Atlas (WTA) panel (18,000 genes) with the NanoString GeoMx digital spatial profiling technology. The TME regions of interest (ROI) were identified using a set of morphology markers consisting of panCK, CD45 and Syto13; 12 ROIs were selected from each sectioned tumor material. The same ROIs were selected at the two testing sites with matching tissue location and morphology characters. Results: GeoMx WTA panel provided an unbiased view of 18,000 protein-coding genes at specific TME regions. These counts were successfully mapped to the 12 ROIs on the tissue. A pseudo-TIS score was generated using the same 18 gene signature from the WTA panel for each ROI. The pseudo-TIS scores from ROIs from the four specimens showed consistency with the specimen-wise TIS score, yet variations were observed across different TME/ROI within each specimen. Whole transcriptome expression results showed high correlation between the two testing sites, providing confidence in the reproducibility of the technology. High resolution fluorescent staining images were able to be fed to downstream quantitative digital analysis to generate complementary spatial data. Citation Format: Caitlin Schroyer, Shanshan Zhou. Spatial transcriptomic study of the tumor microenvironment in HNSCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3652.

Male facial rejuvenation using a combination 633, 830, and 1072nm LED face mask.

Home-based photobiomodulation is a popular treatment modality for patients seeking non-invasive aesthetic treatment. Studies demonstrate that photobiomodulation is effective for skin rejuvenation, which is aimed at improving the overall appearance of the skin by reducing fine lines and wrinkles and improving skin texture, skin tone, and dyspigmentation. Most current skin rejuvenation research focuses on treatments in women. However, men's aesthetics remains an underserved market. A combined red light (RL) and near-infrared (NIR) light-emitting diode (LED) has been designed specifically to target male skin, which may have different physiological and biophysical properties compared to female skin. Herein, the safety and efficacy of a commercially available RL and NIR (633, 830, and 1072nm) LED array designed to be worn as a face mask was assessed. Primary outcomes included adverse events and facial rejuvenation as determined by participant-reported satisfaction scales and quantitative digital skin photography and computer analysis after 6weeks of treatment. The participants reported overall favorable results and improvements in all individual categories, were satisfied with the treatment, and would recommend the product to others. The participants perceived the greatest improvement in fine lines and wrinkles, skin texture, and youthful appearance. Photographic digital analysis demonstrated favorable improvements in wrinkles, UV spots, brown spots, pores, and porphyrins. These results support the use of RL and NIR to treat male skin. Advantages of the LED facemask include its safety, efficacy, convenient home-based use, minimal associated downtime, simple operation, non-invasiveness, and appreciable results in as few as 6weeks.

A Quantitative Digital Analysis of Tissue Immune Components Reveals an Immunosuppressive and Anergic Immune Response with Relevant Prognostic Significance in Glioblastoma.

Objectives: Immunostimulatory therapies using immune checkpoint blockers show clinical activity in a subset of glioblastoma (GBM) patients. Several inhibitory mechanisms play a relevant role in the immune response to GBM. With the objective of analyzing the tumor immune microenvironment and its clinical significance, we quantified several relevant immune biomarkers. Design: We studied 76 primary (non-recurrent) GBMs with sufficient clinical follow-up, including a subgroup of patients treated with a dendritic cell vaccine. The IDH-mutation, EGFR-amplification, and MGMT methylation statuses were determined. Several relevant immune biomarkers, including CD163, CD8, PD1, and PDL1, were quantified in representative selected areas by digital image analysis and semiquantitative evaluation. The percentage of each immune expression was calculated with respect to the total number of tumor cells. Results: All GBMs were wild-type IDH, with a subgroup of classical GBMs according to the EGFR amplification (44%). Morphologically, CD163 immunostained microglia and intratumor clusters of macrophages were observed. A significant direct correlation was found between the expression of CD8 and the mechanisms of lymphocyte immunosuppression, in such a way that higher values of CD8 were directly associated with higher values of CD163 (p < 0.001), PDL1 (0.026), and PD1 (0.007). In a multivariate analysis, high expressions of CD8+ (HR = 2.05, 95%CI (1.02–4.13), p = 0.034) and CD163+ cells (HR 2.50, 95%CI (1.29–4.85), p = 0.007), were associated with shorter survival durations. The expression of immune biomarkers was higher in the non-classical (non-EGFR amplified tumors) GBMs. Other relevant prognostic factors were age, receipt of the dendritic cell vaccine, and MGMT methylation status. Conclusions: In accordance with the inverse correlation between CD8 and survival and the direct correlation between effector cells and CD163 macrophages and immune-checkpoint expression, we postulate that CD8 infiltration could be placed in a state of anergy or lymphocytic inefficient activity. Furthermore, the significant inverse correlation between CD163 tissue concentration and survival explains the relevance of this type of immune cell when creating a strong immunosuppressive environment. This information may potentially be used to support the selection of patients for immunotherapy.

Ultrasound image analysis for detection of fatty liver and grading its severity in buffaloes at Mosul, Iraq

The study aimed to (ⅰ) examines whether quantitative and qualitative digital analysis of hepatic ultrasound images can be used to detect fatty liver and grading its severity, (ⅱ) to estimate the concentration of serum β-hydroxybutyrate content in normal and diseased animals. A total of 50 buffaloes (15 clinically normal and 35 suffered from decreased milk production, loss of appetite, and loss of body weight) 2-4 weeks after parturition were studied. Buffaloes were examined by real-time ultrasound using a 3.5 MHz convex probe. Ultrasound images were saved for qualitative and quantitative analysis, using computer aid Fatty-Liver software (programmed in Matlab language). Based on the digital analysis of ultrasound images, the diseased buffaloes were classified as mild (n=20), moderate (n=7) and severe (n=8) fatty liver affection. Results indicated that the concentration of β-hydroxybutyrate estimated via ELISA were 0.409, 0.735, 0.923, 1.708 mmol/dL in normal, mild, moderate, and severe diseased buffaloes respectively. However, a significant difference has been encountered among normal and diseased animals. It has been concluded that buffaloes at Mosul suffered from fatty liver and digital analysis of ultrasound images was a suitable method for clinical diagnosis, furthermore, estimation of β-hydroxybutyrate was a suitable method for laboratory diagnosis.

Leksiskās daudzveidības kvantitatīvā analīze latviešu prozas izpētē

The notions of richness of language and richness of the vocabulary are characteristics that are often mentioned in the context of practical literary criticism, however, from the theoretical standpoint, the precise meaning of these terms is not clear. Considering that this area of research is not well known in Latvia, the paper first outlines the field of study of vocabulary richness and lexical diversity and maps out the challenges that are related to it, including issues pertaining to selecting the focus and methodology of the study, selecting data, and evaluating the quality of data. Furthermore, the interpretation of the results of the analysis of lexical diversity is discussed, arguing that the vocabulary richness of a literary work does not necessarily indicate only the level of language competencies and artistic skill, it can also be interpreted as a trait of a literary genre or a subject of the artistic choice. Two methods of accounting for lexical diversity are analysed more closely – mean segmental type-token ratio (MSTTR) and moving window type-token ratio distribution (MWTTRD); the main idea and application, as well the place of these approaches among other methods are explained. To conduct the test study, the authors are using a small corpus that entails six Latvian prose writers – Alberts Bels, Augusts Deglavs, Regīna Ezera, Jānis Jaunsudrabiņš, Ivande Kaija, and Zigmunds Skujiņš. In the course of the study it is established that higher values of lexical diversity are achieved by the authors who represent more recent generation – Skujiņš, Ezera, Bels. To suggest further possibilities of employment of these methods in the studies of the stylistics of literary prose, the authors of the paper turn attention towards the diversity of adjectives in the works of the corpus. The MSTTR and MWTTDR of adjectives are measured; the percentage of adjectives that are shared by all authors and the percentage of unique adjectives for each author are being calculated. In conclusion, further developments of quantitative and qualitative digital analysis of prose texts are being suggested.

Could automated analysis of chest X-rays detect early bronchiectasis in children?

Non-cystic fibrosis bronchiectasis is increasingly described in the paediatric population. While diagnosis is by high-resolution chest computed tomography (CT), chest X-rays (CXRs) remain a first-line investigation. CXRs are currently insensitive in their detection of bronchiectasis. We aim to determine if quantitative digital analysis allows CT features of bronchiectasis to be detected in contemporaneously taken CXRs. Regions of radiologically (A) normal, (B) severe bronchiectasis, (C) mild airway dilation and (D) other parenchymal abnormalities were identified in CT and mapped to corresponding CXR. An artificial neural network (ANN) algorithm was used to characterise regions of classes A, B, C and D. The algorithm was then tested in 13 subjects and compared to CT scan features. Structural changes in CT were reflected in CXR, including mild airway dilation. The areas under the receiver operator curve for ANN feature detection were 0.74 (class A), 0.71 (class B), 0.76 (class C) and 0.86 (class D). CXR analysis identified CT measures of abnormality with a better correlation than standard radiological scoring at the 99% confidence level.Conclusion: Regional abnormalities can be detected by digital analysis of CXR, which may provide a low-cost and readily available tool to indicate the need for diagnostic CT and for ongoing disease monitoring. What is Known:• Bronchiectasis is a severe chronic respiratory disorder increasingly recognised in paediatric populations.• Diagnostic computed tomography imaging is often requested only after several chest X-ray investigations.What is New:• We show that a digital analysis of chest X-ray could provide more accurate identification of bronchiectasis features.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00431-021-04061-8.

Knockout of the HMG domain of the porcine SRY gene causes sex reversal in gene-edited pigs

The sex-determining region on the Y chromosome (SRY) is thought to be the central genetic element of male sex development in mammals. Pathogenic modifications within the SRY gene are associated with a male-to-female sex reversal syndrome in humans and other mammalian species, including rabbits and mice. However, the underlying mechanisms are largely unknown. To understand the biological function of the SRY gene, a site-directed mutational analysis is required to investigate associated phenotypic changes at the molecular, cellular, and morphological level. Here, we successfully generated a knockout of the porcine SRY gene by microinjection of two CRISPR-Cas ribonucleoproteins, targeting the centrally located "high mobility group" (HMG), followed by a frameshift mutation of the downstream SRY sequence. This resulted in the development of genetically male (XY) pigs with complete external and internal female genitalia, which, however, were significantly smaller than in 9-mo-old age-matched control females. Quantitative digital PCR analysis revealed a duplication of the SRY locus in Landrace pigs similar to the known palindromic duplication in Duroc breeds. Our study demonstrates the central role of the HMG domain in the SRY gene in male porcine sex determination. This proof-of-principle study could assist in solving the problem of sex preference in agriculture to improve animal welfare. Moreover, it establishes a large animal model that is more comparable to humans with regard to genetics, physiology, and anatomy, which is pivotal for longitudinal studies to unravel mammalian sex determination and relevant for the development of new interventions for human sex development disorders.

Revisiting growth and decline of late Quaternary mega-lakes across the south-central Tibetan Plateau

Lake basins record critical interactions between the hydrosphere, cryosphere and atmosphere. Their sediments and shoreline landforms archive timing, duration, and intensity of past climatic variability and environmental impacts over a variety of timescales. Thousands of lake systems spread across the internally drained Tibetan Plateau today, of which many are only the salty remains of much more expansive paleolakes in the past. This study presents new shoreline ages integrated with quantitative digital topographic analysis and regional geomorphic evidence to reconstruct the sizes and extents of late Quaternary rift lake systems for the south-central Tibetan Plateau. This study presents optically stimulated luminescence (OSL) and the first K-feldspar post-infrared infrared (pIRIR) stimulated luminescence ages for Zabuye Caka and Dawa Tso and tests if these lakes were once part of a singular Pleistocene mega-lake established across four N–S rift systems. Our new results show that two large but separate paleolake systems developed in the latest Pleistocene-earliest Holocene and no lake larger than ∼6460 km2 has existed since at least 41 ka in the Lunggar region. Early Holocene paleolakes expanded up to 7× modern (+220–335 km3) with two to four-fold asymmetric lake expansion in the west compared to the east. Findings corroborate earlier investigations, implying that dramatic runoff increase and/or evapotranspiration reduction prompted rapid earliest Holocene lake expansion. Precipitation variability, in combination with permafrost degradation, temperature and wind strength, governed paleolake moisture balance. Open-system lake behavior and evolving drainage configurations across complex topography explain delayed onsets of lake regression and punctuated lake decline. This suggests abrupt Holocene climate shifts are not necessarily required for punctuated lake decline. Lastly, unique geomorphological and paleoclimatic similarities between Tibetan and East African rift lakes highlight the potential for some Tibetan paleolakes to function as natural climate amplifiers over both short and geologic timescales.

Quantitative digital imaging analysis of HER2 immunohistochemistry predicts the response to anti-HER2 neoadjuvant chemotherapy in HER2-positive breast carcinoma.

Patients with HER2-positive breast cancer commonly receive anti-HER2 neoadjuvant chemotherapy and pathologic complete response (pCR) can be achieved in up to half of the patients. HER2 protein expression detected by immunohistochemistry (IHC) can be quantified using digital imaging analysis (DIA) as a value of membranous connectivity. We aimed to investigate the association HER2 IHC DIA quantitative results with response to anti-HER2 neoadjuvant chemotherapy. Digitized HER2 IHC whole slide images were analyzed using Visiopharm HER2-CONNECT to obtain quantitative HER2 membranous connectivity from a cohort of 153 HER2+ invasive breast carcinoma cases treated with anti-HER2 neoadjuvant chemotherapy (NAC). HER2 connectivity and other factors including age, histologic grade, ER, PR, and HER2 fluorescence in situ hybridization (FISH) were analyzed for association with the response to anti-HER2 NAC. Eighty-three cases (54.2%) had pCR, while 70 (45.8%) showed residual tumor. Younger age, negative ER/PR, higher HER2 DIA connectivity, higher HER2 FISH ratio and copy number were significantly associated with pCR in univariate analysis. Multivariate analysis demonstrated only age, HER2 DIA connectivity, PR negativity, and HER2 copy number was significantly associated with pCR, whereas HER2 DIA connectivity had the strongest association. HER2 IHC DIA connectivity is the most important factor predicting pCR to anti-HER2 neoadjuvant chemotherapy in patients with HER2-positive breast cancer.

CD163+ immune cell infiltrates and presence of CD54+ microvessels are prognostic markers for patients with embryonal rhabdomyosarcoma

Rhabdomyosarcomas (RMS) are rare and often lethal diseases. It is assumed that the tumor microenvironment (TME) of RMS exerts an immunosuppressive function, but there is currently no systematic analysis of the immune cells infiltrating sarcoma tissue. Focusing on two common types of RMS (alveolar [RMA] and embryonal [RME]), we performed a comprehensive immunohistochemical analysis of tumor-infiltrating immune cells in the TME. We performed a qualitative estimation of infiltrating immune cells in the tumor microenvironment by an experienced pathologist as well as a quantitative digital pathology analysis. We found that (1) manual and automatic quantification of tumor-infiltrating immune cells were consistent; (2) RME tumors showed a higher degree of immune cell infiltration than RMA tumors but (3) the number of tumor infiltrating lymphocytes was low compared to other solid tumor types; (4) microvascular density correlated with immune cell infiltration and (5) CD163 positive macrophages as well as CD54 positive microvessels were more often detected in RME than in RMA and correlated with patient overall and event free survival. Our systematic analysis provides a comprehensive view of the immune landscape of RMS which needs to be taken into account for developing immunotherapies for this rare type of cancer.

Variation in the Venom of Parasitic Wasps, Drift, or Selection? Insights From a Multivariate QST Analysis

Differentiation of traits among populations can evolve by drift when gene flow is low relative to drift or selection when there are different local optima in each population (heterogeneous selection), whereas homogeneous selection tends to prevent evolution of such a differentiation. Analyses of geographical variations in venom composition have been done in several taxa such as wasps, spiders, scorpions, cone snails and snakes, but surprisingly never in parasitoid wasps, although their venom should constrain their ability to succeed on locally available hosts. Such a study is now facilitated by the development of an accurate method (quantitative digital analysis) that allows analyzing the quantitative variation of large sets of proteins from several individuals. This method was used here to analyse the venom-based differentiation of four samples of Leptopilina boulardi and five samples of L. heterotoma from populations along a 300 km long south-north gradient in the Rhone-Saone valley (South-East of France). A major result is that the composition of the venom allows to differentiate the populations studied even when separated by few kilometers. We further analyzed these differentiations on the populations (reared under similar conditions to exclude environmental variance) with a QST analysis which compared the variance of a quantitative trait (Q) among the subpopulations (S) to the total variance (T). We also used random forest clustering analyses to detect the venom components the most likely to be adapted locally. The signature of the natural selection was strong for L. heterotoma and L. boulardi. For the latter, the comparison with the differentiation observed at some neutral markers revealed that differentiation was partly due to some local adaptation. The combination of methods used here appears to be a powerful framework for population proteomics and for the study of eco-evolutionary feedbacks between proteomic level and population and ecosystem levels. This is of interest not only for studying field evolution at an intermediate level between the genome and phenotypes, or for understanding the role of evolution in chemical ecology, but also for more applied issues in biological control.

Prostate Cancer: A Correlative Study of Multiparametric MR Imaging and Digital Histopathology.

Purpose To correlate multiparametric magnetic resonance (MR) imaging and quantitative digital histopathologic analysis (DHA) of the prostate. Materials and Methods This retrospective study was approved by the local institutional review board and was HIPAA compliant. Forty patients (median age, 60 years; age range, 44-71 years) who underwent prostate MR imaging consisting of T2-weighted and diffusion-weighted (DW) MR imaging along with subsequent robot-assisted radical prostatectomy gave informed consent to be included. Whole-mount tissue specimens were obtained with a patient-specific mold, and DHA was performed to assess the lumen, epithelium, stroma, and epithelial nucleus. These DHA images were registered with MR images and were correlated on a per-voxel basis. The relationship between MR imaging and DHA was assessed by using a linear mixed-effects model and the Pearson correlation coefficient. Results T2-weighted MR imaging, apparent diffusion coefficient (ADC) of DW imaging, and high-b-value DW imaging were significantly related to specific DHA parameters (P < .01). For instance, lumen density (ie, the percentage area of tissue components) was associated with T2-weighted MR imaging (slope = 0.36 ± 0.05 [standard error], γ = 0.35), ADC (slope = 0.47 ± 0.05, γ = 0.50), and high-b-value DW imaging (slope = -0.44 ± 0.05, γ = -0.44). Differences between regions harboring benign tissue and those harboring malignant tissue were observed at MR imaging and DHA (P < .01). Gleason score was significantly associated with MR imaging and DHA parameters (P < .05). For example, it was positively related to high-b-value DW imaging (slope = 0.21 ± 0.16, γ = 0.18) and negatively related to lumen density (slope = -0.19 ± 0.18, γ = -0.35). Conclusion Overall, significant associations were observed between MR imaging and DHA, regardless of prostate anatomy. © RSNA, 2017 Online supplemental material is available for this article.

Tumor-associated B cells in cutaneous primary melanoma and improved clinical outcome

B cells often infiltrate the microenvironment of human tumors. B cells can both positively and negatively regulate antitumor immune responses. In several human cancers, higher numbers of CD20(+) TAB are associated with a favorable prognosis, whereas in human primary melanomas, this association is contentious. In this study, we determined the association of TAB numbers in cutaneous primary melanoma tissue samples and patients' overall survival. The CD20 immunohistochemistry on archival nonmetastasized and metastasized cutaneous primary melanoma tissues from 2 independent patient cohorts was performed. One cohort was used in class comparison for metastasis, the most important prognostic factor for overall survival, and the other cohort for a subsequent survival analysis. Survival association was further validated with RNA data from a third independent cohort. Whole tissue sections were read automatically via quantitative digital imaging and analysis. Survival data were analyzed by Cox proportional hazard modeling. We discovered that cutaneous primary melanomas without metastasis contain significantly more TAB than primary melanomas that had metastasized. At time of first diagnosis, a higher number of TAB is associated with a significantly better overall survival in patients with cutaneous primary melanomas of >1 mm Breslow depth. Also, higher CD20/CD19 tumor mRNA levels are correlated with a significantly better overall survival. Thus, our data support TAB numbers as a prognostic biomarker in cutaneous primary melanoma patients with a tumor of >1 mm Breslow depth. For a survey in larger studies, whole tissue section analysis seems to be key to accurate assessment of TAB numbers.

Comparative study between quantitative digital image analysis and fluorescence in situ hybridization of breast cancer equivocal human epidermal growth factor receptors 2 score 2+ cases

Background: Optimization of workflow for breast cancer samples with equivocal human epidermal growth factor receptors 2 (HER2)/neu score 2+ results in routine practice, remains to be a central focus of the on-going efforts to assess HER2 status. According to the College of American Pathologists/American Society of Clinical Oncology guidelines equivocal HER2/neu score 2+ cases are subject for further testing, usually by fluorescence in situ hybridization (FISH) investigations. It still remains on open question, whether quantitative digital image analysis of HER2 immunohistochemistry (IHC) stained slides can assist in further refining the HER2 score 2+. Aim of this Work: To assess utility of quantitative digital analysis of IHC stained slides and compare its performance to FISH in cases of breast cancer with equivocal HER2 score 2+. Materials and Methods: Fifteen specimens (previously diagnosed as breast cancer and was evaluated as HER 2− score 2+) represented the study population. Contemporary new cuts were prepared for re-evaluation of HER2 immunohistochemical studies and FISH examination. All the cases were digitally scanned by iScan (Produced by BioImagene [Now Roche-Ventana]). The IHC signals of HER2 were measured using an automated image analyzing system (MECES, www.Diagnomx.eu/meces). Finally, a comparative study was done between the results of the FISH and the quantitative analysis of the virtual slides. Results: Three out of the 15 cases with equivocal HER2 score 2+, turned out to be positive (3+) by quantitative digital analysis, and 12 were found to be negative in FISH too. Two of these three positive cases proved to be positive with FISH, and only one was negative. Conclusions: Quantitative digital analysis is highly sensitive and relatively specific when compared to FISH in detecting HER2/neu overexpression. Therefore, it represents a potential reliable substitute for FISH in breast cancer cases, which desire further refinement of equivocal IHC results.

Next-generation endomyocardial biopsy: the potential of confocal and super-resolution microscopy.

Confocal laser scanning microscopy and super-resolution microscopy provide high-contrast and high-resolution fluorescent imaging, which has great potential to increase the diagnostic yield of endomyocardial biopsy (EMB). EMB is currently the gold standard for identification of cardiac allograft rejection, myocarditis, and infiltrative and storage diseases. However, standard analysis is dominated by low-contrast bright-field light and electron microscopy (EM); this lack of contrast makes quantification of pathological features difficult. For example, assessment of cardiac allograft rejection relies on subjective grading of H&E histology, which may lead to diagnostic variability between pathologists. This issue could be solved by utilising the high contrast provided by fluorescence methods such as confocal to quantitatively assess the degree of lymphocytic infiltrate. For infiltrative diseases such as amyloidosis, the nanometre resolution provided by EM can be diagnostic in identifying disease-causing fibrils. The recent advent of super-resolution imaging, particularly direct stochastic optical reconstruction microscopy (dSTORM), provides high-contrast imaging at resolution approaching that of EM. Moreover, dSTORM utilises conventional fluorescence dyes allowing for the same structures to be routinely imaged at the cellular scale and then at the nanoscale. The key benefit of these technologies is that the high contrast facilitates quantitative digital analysis and thereby provides a means to robustly assess critical pathological features. Ultimately, this technology has the ability to provide greater accuracy and precision to EMB assessment, which could result in better outcomes for patients.

Finite micro-tab system for load control on a wind turbine

Finite micro-tabs have been investigated experimentally to evaluate the potential for load control on wind turbines. Two dimensional full span, as well as multiple finite tabs of various aspect ratios have been studied on an AH93W174 airfoil at different chord wise positions. A force balance was used to measure the aerodynamic loads. Furthermore, the wake vortex system consisting of the Karman vortex street as well as the tab tip vortices was analyzed with a 12-hole probe and hot wire anemometry. Finally, conventional oil paint as well as a quantitative digital flow analysis technique called SMARTviz were used to visualize the flow around the finite tab configurations.Results have shown that the devices are an effective solution to alleviate the airfoils overall load. The influence of the tab height, tab position as well as the finite tab aspect ratio on the lift and lift to drag ratio have been evaluated. It could be shown, that the lift difference can either be varied by changing the tab height as well as by altering the aspect ratio of the finite tabs. The drag of a two-dimensional flap is directly associated with the vortex street, while in the case of the finite tab, the solidity ratio of the tabs has the strongest effect on the drag. Therefore, the application of a finite tab system showed to improve the lift to drag ratio.

Sodium fluorescein in skull base meningiomas: A technical note

ObjectivesSkull base meningiomas are a neurosurgical challenge due to the involvement of neurovascular structures. In this study, the authors present the first study of the trans-operative use of sodium fluorescein (SF) to enhance skull base meningiomas and perform a quantitative digital analysis of the tumors’ pigmentation. The goal of the study was to observe the SF enhancement of skull base meningiomas. Patients and methodsA prospective, within-subjects study was designed and performed. This study included twelve patients with skull base meningiomas. After an initial dissection, digital pictures were taken before and after systematic injections of SF using the same light-source used for the surgical microscope. These pictures were analyzed with software that calculated the wavelengths of the sodium fluorescein before and after the injection of the dye. ResultsThe meningiomas in the sample included the following types: 1 cavernous sinus, 1 olfactory groove, 3 petroclival, 1 tuberculum sellae, 3 sphenoid wings, 1 anterior clinoid, and 2 temporal floor. The SF enhancement in all tumors was strongly positive. ConclusionsThe low cost, universal availability and safety of SF indicate that this dye should be examined in further studies, and its applications in skull-base meningioma surgeries should be further assessed.

High disturbed flow in stent-implanted region predicts target lesion revascularization after everolimus-eluting stent implantation in hemodialysis

Background: High disturbed coronary blood flow was reported to increase the risk of adverse events. The purpose of this study was to assess predictive values of coronary flow velocity (CFV) and an index of disturbed flow for target lesion revascularization (TLR) after everolimus-eluting stent (EES) implantation in hemodialysis (HD) patients. Methods and results: The study group consisted of 81HD patients with 100 coronary lesions (age 69.9±9.1 years, mean ± SD, 59 male) who underwent successful percutaneous coronary intervention (PCI) with EES implantation. The TIMI frame count method and quantitative digital angiographic analysis were performed based on the post-PCI angiogram. CFV (vessel length/TIMI frame count/15) and an index of disturbed flow (Reynolds number: velocity/diameter/density/viscosity) were measured both throughout the target vessel and in the stent-implanted region. Target lesion revascularization (TLR) occurred in 12 lesions (12.0%) at the follow-up period of 20.9±10.4 months. There were no significant differences between TLR group (12 lesions) and Non-TLR group (88 lesions) in terms of CFV and Reynolds number throughout the target vessel (195.4±66.8mm/sec vs. 159.8±56.6, p=0.17, 153.4±75.4 vs. 119.7±59.4, p=0.08, respectively). However, in the stent-implanted region, CFV and Reynolds number of the TLR group were significantly higher than the Non-TLR group (222.8±70.7mm/sec vs. 160.5±77.2, p=0.006, 186.1±65.7 vs. 132.4±71.2, p=0.01, respectively). Multivariate analysis showed that Reynolds number in the stent-implanted region after EES was an independent predictor of TLR in the HD patients (OR: 1.01, 95% CI: 1.00-1.023, p=0.02). Conclusion: In the stent-implanted region, CFV and Reynolds number were significantly higher in the TLR group than the Non-TLR group. High disturbed flow in stent-implanted region after EES may predict the risk of TLR in HD patients.

Abstract 808: Quantitative digital PCR analysis of cancer gene promoter methylation using low amounts of input DNA.

Abstract Silencing of tumor suppressor and tumor-related gene transcription by hypermethylation at promoter CpG motifs is a significant mechanism at work in human tumorgenesis. Various methods have been developed to analyze DNA methylation levels, both across the genome and at specific loci, in order to discover and interrogate disease relevant loci for methylation-based transcriptional control. Methods able to quantitatively measure differences in DNA methylation between normal and cancer cells provide promising sources for biomarker identification and assessment. Here we report on the use of a methylation assay (Zymo Research's OneStep qMethyl™) and digital PCR counting (RainDrop™ dPCR) to enable quantitative measurements of DNA methylation at specific genomic loci using small amounts of non-bisulfite converted DNA. MGMT promoter methylation measurements are used to illustrate the method's robust quantification of small percent methylation changes using low amounts of input DNA. Zymo Research's OneStep qMethyl™ Kit is used for the detection of region-specific DNA methylation via the selective amplification of methylated cytosines after digestion with methylation-sensitive restriction enzymes. RainDance Technologies’ RainDrop™ dPCR system is a highly sensitive tool for precise quantification of nucleic acids using probe-based qPCR reagents. RainDrop offers unique analytical advantages for clinical diagnostics due to its exceptionally high sensitivity for absolute quantification and capability to multiplex assays, using a wide dynamic range of input DNA in a contamination-free and simple workflow. The combination of both technologies enables a novel, fast, and robust method for quantitative methylation analysis of small amounts of DNA. The DNA repair protein O6-methylguanine DNA methyltransferase (MGMT) removes mutagenic alkyl adducts from DNA, and its epigenetic silencing by promoter methylation confers susceptibility to DNA-damaging alkylating agents in glioblastomas, melanomas, and lung cancer cell lines, and is increased in cancers including advanced gastric carcinomas, colorectal tumors, and triple-negative breast cancers. A sensitive, quick, and cost-effective quantitative method for assessing MGMT promoter methylation would greatly enhance the diagnostic potential for this candidate cancer biomarker. Here we show digital quantification of MGMT promoter methylation across the range from 0 to 100%, demonstrating a linear correlation that enables less than 15% methylation changes to be robustly measured using less than 20 nanograms of input DNA. This method provides a sensitive count of region-specific methylation percentages using an easy workflow. Continuing work will make use of RainDrop's unique ability to multiplex assays to enable combining MGMT methylation analysis with additional cancer relevant methylation biomarkers in a single assay. Citation Format: Michael L. Samuels, Ryan Kemp, Frances Long, Jill Petrisko, Lam Nguyen, Manuel Krispin. Quantitative digital PCR analysis of cancer gene promoter methylation using low amounts of input DNA. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 808. doi:10.1158/1538-7445.AM2013-808