本研究主要在於設計一種新式生物反應器，並應用於生產豬瘟病毒疫苗。首先根據所培養細胞的生長特性與原有生物反應器之缺點，改良成新式的生物反應器，並評估此新式生物反應器適用性、效能，以及所培養豬腎細胞之生長代謝情形與豬瘟病毒力價。整個實驗過程大致分為兩個部分，第一個部分探討細胞固定化培養之最適化培養條件與生長代謝情形，第二個部分探討豬瘟病毒培養之最適化培養條件與生長代謝情形。實驗結果發現豬腎細胞(PK-15)以批次方式培養於新式生物反應器，搭配著FIBRA-CEL®載體，成功的進行擴大培養，豬腎細胞最高的生長量達到2.29×109cells/300mL的細胞量。因此，改良之新式生物反應器可提供細胞優越的生長環境，具有擴大規模培養之潛力，可藉由此簡單設備、操作容易、成本低且低能源消耗之新式生物反應器達成細胞製品之生產基座。 / In this study, the production of PK-15 cell using immobilized animal cell culture in a novel bioreactor was investigated.We evaluated the serviceability and efficiency of a design-improved novel bioreactor for the growth and metabolic states of cultured PK-15 cells and the production of HC virus. The entire experiment includes two major stages: (1) investigation of the optimal conditions and metabolic states for the growth of immobilized cells, (2) investigation of the optimal conditions for the production of HC virus. Our results showed that immobilized PK-15 cells on the fibra-cell carries in the design-improved novel bioreactor exhibited their best growth of 2.29×109 cells/300mL.The immobilized conditions for cell culture, can provide a shearing stress of growth state, easy separation of cells from the culture mediu, and a operation of continuously feeding medium, leading to possibility growth of the high density cell and a long period of production;as a result, the efficiency of producing process is promoted. Here,our design-improved novel bioreactor is expected to provide an optimal growth environment of both the cells and viruses for the production of high-yielded, stable, and consistent cellular biological preparations. Furthermore, it will also provide the basis for the production of cell products with advantages of simple-equipped, easy-to-operate, low cost, and low energy consumption. / 致謝 i 中文摘要 ------------------------------------------------------------------------ ii 英文摘要 ------------------------------------------------------------------------ iii 目錄 ------------------------------------------------------------------------ iv 表目錄 ------------------------------------------------------------------------ vi 圖目錄 ------------------------------------------------------------------------ vii 第一章 緒論------------------------------------------------------------------ 1 第二章 文獻回顧------------------------------------------------------------ 3 2-1 豬腎傳代細胞(PK-15 cell) -------------------------------------- 3 2-2 豬瘟病毒------------------------------------------------------------ 4 2-2.1 豬瘟之歷史背景--------------------------------------------------- 4 2-2.2 豬瘟病毒之特性--------------------------------------------------- 7 2-2.3 豬瘟發生原因之探討--------------------------------------------- 11 2-3 生物反應器--------------------------------------------------------- 12 第三章 實驗材料與方法--------------------------------------------------- 21 3-1 細胞------------------------------------------------------------------ 21 3-2 細胞繼代培養------------------------------------------------------ 21 3-3 細胞冷凍保存------------------------------------------------------ 22 3-4 解凍細胞培養------------------------------------------------------ 22 3-5 病毒感染------------------------------------------------------------ 23 3-6 收集病毒------------------------------------------------------------ 24 3-7 豬瘟病毒力價測試------------------------------------------------ 24 3-8 細胞滾瓶培養------------------------------------------------------ 26 3-9 生物反應器操作--------------------------------------------------- 27 3-10 載體上細胞數的測定--------------------------------------------- 33 3-11 葡萄糖的測定------------------------------------------------------ 33 3-12 培養過程中pH值測定------------------------------------------- 34 第四章 結果與討論--------------------------------------------------------- 35 4-1 測試細胞貼附的材料--------------------------------------------- 35 4-2 細胞固定時間的比較--------------------------------------------- 36 4-3 測試不同比例的載體量培養豬腎細胞------------------------ 37 4-4 測試不同接種量--------------------------------------------------- 40 4-5 測試培養基流速對豬腎細胞生長的影響--------------------- 44 4-6 測試培養基停留於培養槽時間對豬腎細胞生長的影響--- 45 4-7 測試豬腎細胞暴露空氣時間對於生長的影響--------------- 47 4-8 測試Bellocell培養豬腎細胞(PK-15)可行性----------------- 49 4-9 測試利用新式生物反應器培樣豬瘟病毒--------------------- 50 第五章 結論與建議--------------------------------------------------------- 53 參考文獻 ------------------------------------------------------------------------ 55 表目錄 表1. 兔化豬瘟疫苗與組織培養豬瘟疫苗的比較------------------ 6 表2. 急性、慢性與遲發型豬瘟比較---------------------------------- 10 表3. .Growth of Various cell Lines in bellocell-500----------------- 18 表4. Comparison of SF-9 cell Growth and BEV production in Various Laboratory bioreators------------------------------------ 19 表5. Comparison of HEK293 Cell growth and Receptor X production in Cell Factory®/20 roller bottles and BelloCell-500Bioreactor------------------------------------------ 20 表6. Reed-Muench Methods法計算方法----------------------------- 26 表7. 比較不同材料培養PK-15 cell所用的載體量---------------- 50 表8. 細胞固定時間的比較所接細胞量與載體量------------------ 51 圖目錄 圖1. Liau提出以潮汐生物反應器圖--------------------------------- 17 圖2. Operation principle of Bellocell system------------------------- 18 圖3. 新式生物反應器(novel reactor)-潮汐式生物反應器(tidal typereactor)之運作流程圖--------------------------------------- 30 圖4. 比較不同材料培養PK-15 cell ---------------------------------- 63 圖5. 比較不同時間細胞的貼附量------------------------------------ 64 圖6. 測試的不同比例載體量培養豬腎細胞生長曲線------------ 65 圖7. 測試的不同比例載體量培養豬腎細胞培養過程glucose消耗趨勢------------------------------------------------------------ 66 圖8. 測試的不同比例載體量培養豬腎細胞培養過程pH變化------------------------------------------------------------------------ 67 圖9. 測試不同接細胞量培養在10g carrier生長曲線------------- 68 圖10. 測試不同接細胞量培養在10g 載體glucose消耗趨勢----- 69 圖11. 測試不同接細胞量培養在10g carrier pH趨勢--------------- 70 圖12. 測試流速對豬腎細胞生長的影響------------------------------ 71 圖13. 測試流速對細胞影響的葡萄糖消耗--------------------------- 72 圖14. 測試流速對豬腎細胞生長影響pH值-------------------------- 73 圖15. 測試培養基停留時間對豬腎細胞生長影響------------------ 74 圖16. 測試培養基holding時間對豬腎細胞生長的影響之葡萄糖趨勢--------------------------------------------------------------- 75 圖17. 測試培養基holding時間對細胞的影響之pH值趨勢------- 76 圖18. 測試豬腎細胞暴露空氣時間對生長的影響------------------ 77 圖19. 測試豬腎細胞暴露空氣對生長的影響葡萄糖消耗趨勢--- 78 圖20. 測試豬腎細胞暴露空氣對細胞生長的影養pH值趨勢---- 79 圖21. Bellocell反應器培養豬腎細胞---------------------------------- 80 圖22. Bellocell培養豬腎細胞葡萄糖消耗趨勢---------------------- 81 圖23. Bellocell培養豬腎細胞pH值趨勢----------------------------- 82 圖24. 測試利用新式生物反應器培養豬瘟病------------------------ 83

This thesis presents a novel Dynamic Gesture Language Recognition (DGLR) system for human-robot and inter-robot communication. We developed and implemented an experimental setup consisting of a humanoid robot/android able to recognize and execute in real time all the arm gestures of the Dynamic Gesture Language (DGL) in similar way as humans do. Our DGLR system comprises two main subsystems: an image processing (IP) module and a linguistic recognition system (LRS) module. The IP module enables recognizing individual DGL gestures. In this module, we use the bag-of-features (BOFs) and a local part model approach for dynamic gesture recognition from images. Dynamic gesture classification is conducted using the BOFs and nonlinear support-vector-machine (SVM) methods. The multiscale local part model preserves the temporal context. The IP module was tested using two databases, one consisting of images of a human performing a series of dynamic arm gestures under different environmental conditions and a second database consisting of images of an android performing the same series of arm gestures. The linguistic recognition system (LRS) module uses a novel formal grammar approach to accept DGL-wise valid sequences of dynamic gestures and reject invalid ones. LRS consists of two subsystems: one using a Linear Formal Grammar (LFG) to derive the valid sequence of dynamic gestures and another using a Stochastic Linear Formal Grammar (SLFG) to occasionally recover gestures that were unrecognized by the IP module. Experimental results have shown that the DGLR system had a slightly better overall performance when recognizing gestures made by a human subject (98.92% recognition rate) than those made by the android (97.42% recognition rate).

This master's thesis focuses on smartphone game for iOS, which uses recognition of face features and other information, which can be obtained from a smartphone's camera and sensors. This work describes a few approaches for real-time face detection and then introduces and compares possibilities for such task on iOS. Moreover, the thesis contains a draft of the final game and its levels. The game showcases various technologies in its levels such as object detection, processing an image color and others. Finally, the thesis introduces the final form of the game that is released and available on the App Store.

This paper is focused on object learning and recognizing in the image and in the image stream. More specifically on learning and recognizing humans or theirs parts in case they are partly occluded, with possible usage on robotic platforms. This task is based on features called Histogram of Oriented Gradients (HOG) which can work quite well with different poses the human can be in. The human is split into several parts and those parts are detected individually. Then a system of voting is introduced in which detected parts votes for the final positions of found people. For training the detector a linear SVM is used. Then the Kalman filter is used for stabilization of the detector in case of detecting from image stream.

This work focuses on classification and recognition of traffic signs in image. It describes briefly some used methods a deeply describes chosen system including extensions and method for creating models needed for classification. There's described implementation of library and demonstration program including important pieces of knowledge discovered during development. There're also results of some experiments and possible enhancements in conclusion.

Unmanned Aerial Vehicles (UAVs) provides a platform for many automated tasks and with an ever increasing advances in computing, these tasks can be more complex. The use of UAVs is expanded in this thesis with the goal of Search and Rescue (SAR), where a UAV can assist fast responders to search for a lost person and relay possible search areas back to SAR teams. To identify a person from an aerial perspective, low-level Histogram of Oriented Gradients (HOG) feature descriptors are used over a segmented region, provided from thermal data, to increase classification speed. This thesis also introduces a dataset to support a Bird’s-Eye-View (BEV) perspective and tests the viability of low level HOG feature descriptors on this dataset. The low-level feature descriptors are known as Boosted Histogram of Oriented Gradients (BHOG) features, which discretizes gradients over varying sized cells and blocks that are trained with a Cascaded Gentle AdaBoost Classifier using our compiled BEV dataset. The classification is supported by multiple sensing modes with color and thermal videos to increase classification speed. The thermal video is segmented to indicate any Region of Interest (ROI) that are mapped to the color video where classification occurs. The ROI decreases classification time needed for the aerial platform by eliminating a per-frame sliding window. Testing reveals that with the use of only color data iv and a classifier trained for a profile of a person, there is an average recall of 78%, while the thermal detection results with an average recall of 76%. However, there is a speed up of 2 with a video of 240x320 resolution. The BEV testing reveals that higher resolutions are favored with a recall rate of 71% using BHOG features, and 92% using Haar-Features. In the lower resolution BEV testing, the recall rates are 42% and 55%, for BHOG and Haar-Features, respectively.

Information spielt in der Natur eine zentrale Rolle. Als intrinsischer Teil des genetischen Codes ist sie das Grundgerüst jeder Struktur und ihrer Entwicklung. Im Speziellen dient sie auch Organismen, ihre Umgebung wahrzunehmen und sich daran anzupassen. Die Grundvoraussetzung dafür ist, dass sie Information ihrer Umgebung sowohl messen als auch interpretieren können, wozu Zellen komplexe Signaltransduktionswege entwickelt haben. In dieser Arbeit konzentrieren wir uns auf Signalprozesse in S.cerevisiae die von osmotischem Stress (High Osmolarity Glycerol (HOG) Signalweg) und der Stimulation mit α-Faktor (Pheromon Signalweg) angesprochen werden. Wir wenden stochastische Modelle an, die das intrinsische Rauschen biologischer Prozesse darstellen können, um verstehen zu können wie Signalwege die ihnen zur Verfügung stehende Information umsetzen. Informationsübertragung wird dabei mit einem Ansatz aus Shannons Informationstheorie gemessen, indem wir sie als einen Kanal in diesem Sinne auffassen. Wir verwenden das Maß der Kanalkapazität, um die Genauigkeit des Phosphorelays einschränken zu können. In diesem Modell, simuliert mit dem Gillespie Algorithmus, können wir durch die Analyse des Signalverhaltens den Parameterraum zusätzlich stark einschränken. Eine weitere Herangehensweise der Signalverarbeitung beschäftigt sich mit dem “Crosstalk” zwischen HOG und Pheromon Signalweg. Wir zeigen, dass die Kontrolle der Signalspezifizität vor allem bei Scaffold-Proteinen liegt, die Komponenten der Signalkaskade binden. Diese konservierten Motive zellulärer Signaltransduktion besitzen eine geeignete Struktur, um Information getreu übertragen zu können. Im letzten Teil der Arbeit untersuchen wir potentielle Gründe für die evolutionäre Selektion von Scaffolds. Wir zeigen, dass ihnen bereits durch die Struktur des Mechanismus möglich ist, Informationsgenauigkeit zu verbessern und einer verteilten Informationsweiterleitung sowohl dadurch als auch durch ihre Robustheit überlegen sind. / Information plays a ubiquitous role in nature. It provides the basis for structure and development, as it is inherent part of the genetic code. It also enables organisms to make sense of their environments and react accordingly. For this, a cellular interpretation of information is needed. Cells have developed sophisticated signaling mechanisms to fulfill this task and integrate many different external cues with their help. Here we focus on signaling that senses osmotic stress (High Osmolarity Glycerol (HOG) pathway) as well as α-factor stimulation (pheromone pathway) in S.cerevisiae. We employ stochastic modeling to simulates the inherent noisy nature of biological processes to assess how systems process the information they receive. This information transmission is evaluated with an information theoretic approach by interpreting signal transduction as a transmission channel in the sense of Shannon. We use channel capacity to both constrain as well as quantify the fidelity in the phosphorelay system of the HOG pathway. In this model, simulated with the Gillespie Algorithm, the analysis of signaling behavior allows us to constrain the possible parameter sets for the system severely. A further approach to signal processing is concerned with the mechanisms that conduct crosstalk between the HOG and the pheromone pathway. We find that the control for signal specificity lies especially with the scaffold proteins that tether signaling components and facilitate signaling by trans-location to the membrane and shielding against miss-activation. As conserved motifs of cellular signal transmission, these scaffold proteins show a particularly well suited structure for accurate information transmission. In the last part of this thesis, we examine the potential reasons for an evolutionary selection of the scaffolding structure. We show that due to its structure, scaffolds are increasing information transmission fidelity and outperform a distributed signal in this regard.

Manure contains nutrients for crop growth; however, overapplication, with time, can result in excess nutrients in soil, which can subsequently be lost in surface runoff. <p>The general purpose of this research is to study the effect of liquid hog manure, applied as an agricultural fertilizer, on water chemistry of surface runoff from rainfall simulation tests. Specifically the research focuses on runoff water chemistry comparisons between lands receiving hog manure at different rates, via different injection methods, and upon different slope positions. <p>To examine these objectives, soil nutrient supply rates (P, NH4-N, and NO3-N) of the 0 5 cm depth of soil adjacent to rainfall simulation positions, and runoff water chemistry (TP, OP, NH4-N, NO3-N, DOC, Cl- and coliforms) during rainfall simulation tests were collected before and after manure addition. <p> Generally, manure application did increase soil NH4-N and NO3-N supply rates, and runoff NH4-N concentration. Soil P supply rate and runoff TP concentration were not affected by the manure addition; however, runoff OP concentration at one site (Perdue) increased significantly due to manure addition. The manure treatments applied in this study did not cause any significant increases in fecal or total coliform in runoff from rainfall simulation tests conducted 7 8 months after manure application. None of the water quality parameters exceeded the Guidelines for Canadian Drinking Water Quality. <p> Manure injection method (regular versus low soil surface disturbance) had consistent effects on runoff chemistry, but application rate did not. The regular disturbance method had significantly higher concentrations of water quality parameters than the low disturbance method. <p> The position of the test on the slope did not result in any consistent trends in runoff chemistry, whether before or after manure addition. Foot slope positions had higher soil NH4-N supply rates than upper slope positions, both before and after manure addition. Soil NH4-N, NO3-N, and P supply rates between landscape positions were not likely influenced by manure addition. <p> Regression tests between soil nutrient supply rates and runoff chemistry indicate that soil NH4-N supply rates are a good index to predict runoff NH4-N concentration, but soil P did not predict runoff P.

Manure contains nutrients for crop growth; however, overapplication, with time, can result in excess nutrients in soil, which can subsequently be lost in surface runoff. <p>The general purpose of this research is to study the effect of liquid hog manure, applied as an agricultural fertilizer, on water chemistry of surface runoff from rainfall simulation tests. Specifically the research focuses on runoff water chemistry comparisons between lands receiving hog manure at different rates, via different injection methods, and upon different slope positions. <p>To examine these objectives, soil nutrient supply rates (P, NH4-N, and NO3-N) of the 0 5 cm depth of soil adjacent to rainfall simulation positions, and runoff water chemistry (TP, OP, NH4-N, NO3-N, DOC, Cl- and coliforms) during rainfall simulation tests were collected before and after manure addition. <p> Generally, manure application did increase soil NH4-N and NO3-N supply rates, and runoff NH4-N concentration. Soil P supply rate and runoff TP concentration were not affected by the manure addition; however, runoff OP concentration at one site (Perdue) increased significantly due to manure addition. The manure treatments applied in this study did not cause any significant increases in fecal or total coliform in runoff from rainfall simulation tests conducted 7 8 months after manure application. None of the water quality parameters exceeded the Guidelines for Canadian Drinking Water Quality. <p> Manure injection method (regular versus low soil surface disturbance) had consistent effects on runoff chemistry, but application rate did not. The regular disturbance method had significantly higher concentrations of water quality parameters than the low disturbance method. <p> The position of the test on the slope did not result in any consistent trends in runoff chemistry, whether before or after manure addition. Foot slope positions had higher soil NH4-N supply rates than upper slope positions, both before and after manure addition. Soil NH4-N, NO3-N, and P supply rates between landscape positions were not likely influenced by manure addition. <p> Regression tests between soil nutrient supply rates and runoff chemistry indicate that soil NH4-N supply rates are a good index to predict runoff NH4-N concentration, but soil P did not predict runoff P.

Computational imaging has been playing an important role in understanding and analysing the captured images. Both image segmentation and restoration has been in-tegral parts of computational imaging. The studies performed in this thesis has been focussed toward developing novel algorithms for image segmentation and restoration. Study related to usage of Morozov Discrepancy Principle in Di use Optical Imaging was also presented here to show that hyper parameter selection could be performed with ease. The Laplacian of Gaussian (LoG) and Canny operators use Gaussian smoothing be-fore applying the derivative operator for edge detection in real images. The LoG kernel was based on second derivative and is highly sensitive to noise when compared to the Canny edge detector. A new edge detection kernel, called as Helmholtz of Gaussian (HoG), which provides higher di suavity is developed in this thesis and it was shown that it is more robust to noise. The formulation of the developed HoG kernel is similar to LoG. It was also shown both theoretically and experimentally that LoG is a special case of HoG. This kernel when used as an edge detector exhibited superior performance compared to LoG, Canny and wavelet based edge detector for the standard test cases both in one- and two-dimensions. The linear inverse problem encountered in restoration of blurred noisy images is typically solved via Tikhonov minimization. The outcome (restored image) of such min-imitation is highly dependent on the choice of regularization parameter. In the absence of prior information about the noise levels in the blurred image, ending this regular-inaction/hyper parameter in an automated way becomes extremely challenging. The available methods like Generalized Cross Validation (GCV) may not yield optimal re-salts in all cases. A novel method that relies on minimal residual method for ending the regularization parameter automatically was proposed here and was systematically compared with the GCV method. It was shown that the proposed method performance was superior to the GCV method in providing high quality restored images in cases where the noise levels are high Di use optical tomography uses near infrared (NIR) light as the probing media to recover the distributions of tissue optical properties with an ability to provide functional information of the tissue under investigation. As NIR light propagation in the tissue is dominated by scattering, the image reconstruction problem (inverse problem) is non-linear and ill-posed, requiring usage of advanced computational methods to compensate this. An automated method for selection of regularization/hyper parameter that incorporates Morozov discrepancy principle(MDP) into the Tikhonov method was proposed and shown to be a promising method for the dynamic Di use Optical Tomography.