Development of a bioengineered transplantable liver graft with re-endothelialized vasculature for generating humanized organ

Kamaleldin Hany Hussein Yousseif 강원대학교 일반대학원 2016 국내박사

Each year, there is a severe shortage of livers for organ transplant. One approach to alleviate this demand is to reconstruct livers using tissue engineered scaffolds. Such scaffolds may be produced through the decellularization of whole organs. Decellularization is a process of cell removal to create a natural extracellular matrix (ECM) for potential transplantation. Ideally, the resulting ECM would retain the organ-specific architecture and chemical composition to guide implanted cells into functional structures and eventually full-functional organs. In the first study, we decellularized the rat liver followed by reseeding using hepatocellular carcinoma cells (HepG2 cells) to optimize the decelularization and recellularization techniques in small-scale and to evaluate if the ECM microenvironment and cues are able to support the cell function and behavior similar to the in vivo state or not. The cell function and tumorgenicity were compared to that HepG2 cells grown as monolayers or on Matrigel. The reseeded scaffolds were infused by antitumor drug (Methotrexate) and the cell response was compared to that HepG2 grown as monolayer. Cells grown in scaffolds seemed to respond to the drug in an analogous manner to its known activity in vivo. These findings strengthen the potential use of rat liver scaffolds for screening HCC drugs. Additionally, this data indicates the ability to produce a whole decellularized liver by perfusion of chemicals through the vasculature of the organ. In the second study, we evaluated the effect of different sterilizing agents on the biological properties of decellularized porcine liver slices as animal-originating scaffolds must undergo treatment to minimize or eliminate the host immune response and to avoid zoonotic disease transmission after in vivo implantation in recipients. Decellularized slices were treated with either PAA or ethanol or slightly acidic electrolyzed water (SAEW), and then DNA and ECM components content were quantified in addition to cell attachment and proliferation. We determined sterilization efficiency by culturing scaffolds in culture medium and on blood agar. PAA and SAEW treatments achieved the highest efficiency of sterilization compared to that of the ethanol treated scaffolds, and were able to remove a considerable fraction of DNA from decellularized livers. The retained glycosaminoglycan content decreased in all treatments in the following order: SAEW, ethanol, and PAA. Ethanol caused a significant loss in collagen content compared to the other groups. SAEW-treated scaffolds supported cell attachment and proliferation at a significantly higher rate than other groups. These data suggest that SAEW is highly efficient for sterilizing scaffolds and allowed the scaffolds to retain their bioactivity in addition to its high efficiency for cell remnant removal. In the third study, we decellularized the right lateral lobe of the porcine liver to evaluate its biocompatibility. Decellularization of porcine livers was achieved by perfusion of detergent-based solutions through the right branch of the portal vein. The resultant organ-shaped structure was spongy and translucent. The decellularized lobe preserved the three-dimensional architecture, ultrastructure, extracellular matrix components, and vasculature. Scaffolds were almost depleted of DNA in addition to antigenic and pathogenic antigens, which are considered barriers to xenotransplantation. The human immune response against scaffolds was considered non-significant. Our matrices were non-cytotoxic and biodegradable. In summary, we successfully developed a biocompatible porcine hepatic lobe for future liver regeneration and bioengineering. In the fourth study, we tried to solve the problem of lack of the intact endothelial layer in the vascular network of decellularized solid organs which can result in blood clotting after transplantation even with administrating anti-coagulation drugs. We hypothesized that heparin-gelatin (HG) mixture can act as an antithrombotic coating reagent in addition to inducing factor for attachment and migration of endothelial cells (ECs) on surfaces of the vascular walls within decellularized porcine liver with subsequent enhancing of parenchymal cell function and viability. We coated portal vein and hepatic artery walls in decellularized liver using HG, then repopulated with EA.hy926 human endothelial cells (ECs) and maintained under a vascular flow through portal vein in a bioreactor for 10 days. Cell location, function, and proliferation were evaluated to confirm the efficiency of re-endothelialization. To assess thrombogenicity of re-endothelialized livers, scaffolds were perfused in ex-vivo system with porcine blood. Then, we co-cultured both HepG2 cells and endothelial cells to check the effect of efficient re-endothelialization on hepatic cells. ECs covered the vascular compartments within decellularized scaffolds with the ability to maintain their function and proliferation. No thrombosis was observed after 24 h blood perfusion in scaffolds precoated by HG in addition; platelets were neither adhered to surface of vessels nor penetrated into parenchyma, indicating a fully endothelialized vascular tree. HepG2 cells displayed a higher function and viability in scaffolds re-endothelialized after precoating by HG compared to scaffolds re-endothelialized without precoating or HepG2 cell-reseeded scaffolds. In vivo transplantation of bioengineered liver scaffolds with/without precoating by HG for 60 min confirmed that HG-precoating treatment can delay the coagulation and enhance the HepG2 cell function. These results provide a proof of principle for the generation of a transplantable liver graft with reconstructed blood vessels as a potential treatment for liver disease in human.

Soil bioengineering 技術을 이용한 山林毁損地 復元 및 安定化에 관한 硏究

이해주 강원대학교 일반대학원 2007 국내박사

The influence of plants on slope stability and natural recover in degraded forest area have been one of the most important subjects of research, especially the role of plant's roots in reinforcing soil strength and recovering methods of degraded forest by plants. The purpose of this study is to evaluate the mechanical effect of roots on preventing the shallow-seated landslide in degraded forest area and to find the methods for natural recovering of degraded forest area by the native plants The Based on the community structural attributes, such as species composition, species diversity, invasion species, in situ surveys on distributions of dominant plants species were carried out to find effective restoration methods in the degraded forest by artificial or naturally disturbances. The results were as follows : In the vegetation occurrence amount in closely relation with early stabilization of degraded area, herbaceous plants were early most dominant in the first year of degrading, but the occurrence number of woody plants tended to be more dominant after second year. While the dominance of herbaceous plants tended to be decreased as years passed, but that of tree plants were increased. Consequently, the effective combination to employ for restoration of the degraded area were Pinus koraiensis + Salix koreensis, Weigela subsessilis + Stephanandra incida, Larix leptolepis + Festuca ovina, Lespedeza bicolor + Eularia speciosa, Larix leptolepis + Aralia elata, Aralia elata + Fragaria ananassa, Aralia elata + Lysimashia clethroides, and Weigela subsessilis + Lysimashia clethroides. The studies were conducted to provide basic data for environmental recovering of mountain lands disturbed by various earthworks, with investigating their germination and growth status after sowing the seeds of the 7 pioneer plant species (woody : Lespedeza bicolor, Pinus densiflora and Rubus crataegifolius and herbaceous : Artemisia princes, Miscanthus sinensis, Themeda triandra var. japonica and Spodiopgon cotulifer) on a disturbed mountain area. After sowing on the 14th of June, 2006, checking on the sowing area was done at an interval of 7 to 15days. In result, mean times to germinate were 21d for Artemisia princes and Themeda triandra, 49d for Lespedeza bicolor and 79d for Pinus densiflora, which are slightly longer than exotic plant species. And the germination percentage were 65.8% of Artemisia princes and 61.3%of Lespedeza bicolor. Although the germination ratiosof the grass species were low, they are favorable to apply to recover the disturbed lands because the seeds are very small. Populations of survival plants after a lapse of one year from sowing were 48 trees/m of Lespedeza bicolor and 44 plants/m of Artemisia princes and land coverage ratios were 46% of Lespedeza bicolor and 38% of the grass species. Therefore the Lespedeza bicolor and grass species are slightly better than the others in the land coverage To evaluate the effects of tree roots on slope stability, characteristics of the root distribution of Pinus koraiensis were investigated and simulated. The total root volume and the number of roots in each 0.5 cm diameter class were investigated by every 10 cm soil layer. Using these results, a simulation model was developed to estimate the quantity and distribution of tree roots at an arbitrary depth. Standard error between measured and simulated value of root distribution was used to evaluate the developed root distribution simulation model that can be estimate the root number at an arbitrary depth. The results shows that measured and simulated distributions of root were similar with the exception of surface 20㎝ soil. To study stabilization effects on restoration of degraded forest area by shrub, the nine species were selected through previous study and in-situ pull-out test was carried out. After pull-out testing, root volume, root collar diameter, root dry weight and height were measured and also correlations with pull-out resistance forces were analyzed. The highest correlation with pull-out resistance was root collar and root volume. Especially, mean values of pull-out resistances by species comes Salix koreensis, Crepidiastrum sonchifolium, Aralia elata, Stephanandra incisa, Zanthoxylum schinifolium and Weigela subsessilis in that order. To evaluate the effects of tree roots on slope stability, characteristics of the root distribution of Pinus koraiensis were investigated and simulated. The total root volume and the number of roots in each 0.5 cm diameter class were investigated by every 10 cm soil layer. Using these results, a simulation model was developed to estimate the quantity and distribution of tree roots at an arbitrary depth. Standard error between measured and simulated value of root distribution was used to evaluate the developed root distribution simulation model that can be estimate the root number at an arbitrary depth. The results shows that measured and simulated distributions of root were similar with the exception of surface 20㎝ soil. Considering mentioned above results, it is possible to evaluate the effects of the roots on preventing shallow-seated landslides. The results shows that the effect of the root reinforcement on slope stability is absolutely important and that natural recovering in degraded forest area should consider root reinforcement to prevent shallow-seated landslides. Also, stabilization methods by the plants were showed through soil bioengineering which is kinds of natural recovering technic. 훼손된 사면의 피해경감 및 예방을 위하여 사면의 안전율을 증진시킬 수 있는 생물학적 복원공법의 실시가 필요하다. 이러한 방법 중 생물학적 복원공법(Soil Bioengineeing)은 자체적으로 재생 가능한 생물을 이용하기 때문에 야생동물의 서식처 및 먹이를 제공하는 등 건전한 생태계를 유지시키며, 또한 경관보호 측면에서도 그 중요성이 크다고 할 수 있다. 또한 수목의 근계는 토양내부의 버팀벽 역할로 토양을 고정하여 사면의 붕괴를 방지하며, 토양의 공극상태를 양호하게 하여 줌으로써 수자원 함양을 증대시킨다. 또한 수목은 증산작용으로 토양내부의 수분을 적절하게 유지시켜 주며, 낙엽 및 지조물 등은 강수의 저장 및 유출수의 흐름을 완화시키는 긍정적인 효과를 거양하고자 수행하였다. 산림훼손지 침입식생의 계량생태학적 연구에서는 산림훼손지에 조기 안정화와 밀접한 관계를 가지고 있는 식생발생량과 간장, 근원경, 수관폭 등 생육상황을 조사하여 중요도, 종다양도, 균재도 등을 산출하여 종합 검토하였으며, 산림훼손지의 복원 가능 수종으로는 싸리나무, 병꽃나무, 산딸기, 산초나무, 버드나무, 국수나무, 두릅나무, 쑥, 고들빼기, 새류 등이 유리한 것으로 나타났으며, 특히 종간 상관관계가 높게 나타난 잣나무+버드나무, 병꽃나무+국수나무, 낙엽송+김의털, 싸리+개억새, 낙엽송+두릅나무, 두릅나무+딸기, 두릅나무+큰까치수영, 병꽃나무+큰까치수영, 국수나무+대사초의 경우 혼파 또는 혼식이 가능할 것으로 사료된다. 산림훼손지 복원종의 종자 발아는 산림훼손지 침입식생의 계량생태학적 연구에서 선발된 10종 중 종자확보가 가능한 7개 종에 대하여 현지발아 시험을 하였으며, 그 외 종자의 발아가 잘되는 종을 선발하여 훼손지에 적용하고자 실내 배지발아시험을 병행하여 수행하였다. 그 결과 60%이상 발아된 싸리, 쑥을 제외한 산딸기, 기름새, 엇새, 솔새, 소나무 등에 대하여는 발아촉진처리 방법 등의 연구가 수행되어야 할 것으로 사료되며, 생립본수는 싸리와 쑥이 가장 좋은 것으로 나타났다. 특히 발아촉진처리를 하지 않은 무처리에서 90%이상 발아된 종으로 가는쑥부쟁이, 쑥부쟁이, 맑은대쑥, 뽕나무, 쉽싸리 등이 사면 복원에 적용이 가능할 것으로 사료된다. 관목류에 의한 훼손지 안정화 연구를 위하여 산림훼손지 칩입식생의 계량생태학적 연구를 통해 선발된 9종을 대상으로 근계에 의한 뿌리의 보강효과를 구명하기 위하여 인발시험을 하였다. 인발시험은 지하부의 뿌리와 자상부의 수간 등의 길이 및 근원경, 체적, 건중량을 측정하여 요인별로 상관관계를 분석한 결과, 인발력과 가장 밀접한 상관을 가지고 있는 요인은 근원경과 체적으로 나타났으며, 뿌리의 인발력은 체적과 근원경에 비례해 인발력이 증가하는 것으로 나타났다. 특히 수종별 평균인발력은 버드나무, 고들빼기, 두릅나무, 국수나무, 산초나무, 병꽃나무, 순으로 나타났다. 교목류에 의한 훼손지 안정화 연구는 수목뿌리가 산지사면 붕괴에 미치는 영향을 파악하기 위해 수목뿌리의 토질강도 보강효과를 현지조사와 실험을 통해 규명하고, 수목근계의 효과를 파악하기 위하여 소나무 근계분포의 특성과 뿌리의 인장전단강도 특성을 중심으로 평가하였으며, 소나무의 뿌리의 분포에 관한 특성 및 규칙성을 조사하고, 뿌리의 분포를 재현할 수 있는 모델을 통해 소나무 뿌리의 지하 분포정보를 파악하였다. 뿌리의 분포상태에 관한 특징 및 규칙성으로서 10cm 토층내의 근의 체적분포는 와이블 분포함수에 의해서 표현하는 것이 가능하며, 각 10㎝ 토층 내의 근의 직경과 본수 비율과 뿌리의 직경과 체적을 대수함수에 의해서 표현할 수 있었다. 특히 위에서 개발되어진 모델에서는 H, DBH, Zmax의 3인자를 입력하면, 깊이별에 직경계급별의 뿌리의 본수가 출력되도록 하였으며, 공시목을 대상으로 한 시뮬레이션 결과에 대한 평가는 뿌리의 저면에서는 비교적 정확한 예측을 하고 있기 때문에 수목이 존재하는 무한장 사면안정 해석에 필요한 붕괴면의 뿌리단면적률을 예측하기 위한 목적을 만족하는 것으로 판단된다. 이상의 결과를 종합하면 훼손된 사면의 피해 예방을 위하여 사면의 안전률을 증진시킬 수 있고 사면의 안전률을 유지시킬 수 있는 생물학적 복원공법을 위하여 우점도, 발아율, 생장량, 뿌리인발력 등을 종합적으로 평가하여 과학적인 검증을 하고 Soil Bioengineeing기술을 통하여 산림훼손지의 안정화 방안을 제시하였다.

Rapid fabrication of 3D printed masters with glassy surfaces of micropatterns for bioengineering applications

Ji Joon Hwan 고려대학교 KU-KIST융합대학원 2024 국내석사

Multifunctional mesoporous silica nanoparticles for immunotherapy

Cha, Bonggeun Sungkyunkwan university 2019 국내박사

The term ‘immune-nano-bioengineering’ is used to describe to design materials, delivery vehicles and molecules both to manipulate and to better understand the immune system. The field is rapidly evolving along with advances in understanding of immunology and is also contributing to knowledge of basic immunology. Over the past decade, particulate carriers have emerged as an attractive means for enhancing the delivery efficacy and potency of vaccines and associated immunomodulatory molecules. Specifically, engineered nanomaterials are being extensively studied for a wide variety of applications. Recently, a lot of researches focus on the immunotherapy for overcoming cancer, chronic viral disease, infection, allergy, and autoimmunity. Along with the development of nanotechnology, these research trends are accomplished progression to immune-nano-bioengineering. To apply nano-engineered materials to immunotherapy is accompanied delivery property. Especially, multi-functional inorganic nanocomposite can be proposed for the application to immunotherapy, subdividing immunoactivation, suppression, and modulation. For example, in case of based on mesoporous silica nanoparticles, changes of physical and chemical properties, different surface engineering, and composition of theragnostic function can affect the immunoengineering properties. Furthermore, both interior and exterior side of mesoporous silica nanoparticles can be decorated with various functional inorganic nanomaterials simultaneously. These multi-functional inorganic nanomaterials become one of the best candidates to apply various immunotherapy areas. In this thesis, the four parts of ‘immune-nano-bioengineeging’ are investigated the synthesis of multifunctional mesoporous silica nanoparticles, included changing of pore structure, immunosuppression through ROS scavenging, immunoactivation by cancer vaccine, and immunomodulation to macrophage polarization. Firstly, a facile method to expand pore size of mesoporous silica nanoparticles embedding superparamagnetic nanocrystals is demonstrated. Uniform extra-large pore mesoporous silica nanoparticles (XL-MSNs) with 30 nm pores were synthesized using organic additives to form extra-large pores in the presence of inorganic seed nanoparticles to generate uniform particles. Furthermore, XL-MSNs show a simple size and pore morphology controlling. Furthermore, XL-MSNs showed significantly higher loading capacity for the model proteins with different molecular weights compared to conventional small pore MSNs. Secondly, the use of XL-MSNs as a cancer vaccine through the delivery of protein antigen and danger signal was demonstrated. Additional surface modification of XL-MSNs resulted in significantly higher loading of antigenic protein and toll-like receptor 9 (TLR9) agonist compared with conventional small-pore MSNs. In vitro culture of bone marrow-derived DCs (BMDCs) in the presence of XL-MSNs loaded with antigen and TLR9 agonist led to enhanced DC activation and antigen presentation and increased secretion of proinflammatory cytokines. An in vivo study demonstrated efficient targeting to draining lymph nodes, induction of antigen-specific CTLs, enhanced suppression of tumor growth after vaccination, and prevention of tumor growth after rechallenge of cancer cells into vaccinated mice due to a significant generation memory T cells. Thirdly, XL-MSNs as delivery carrier of IL-4 for macrophage polarization were demonstrated. XL-MSNs showed significantly higher loading capacity for IL-4 compared to conventional small pore MSNs. XL-MSNs did not induce the secretion of proinflammatory cytokines in vitro or in vivo, which is advantageous for polarization of anti-inflammatory M2 macrophages. Finally, their ability to deliver the M2-polarizing cytokine IL-4 to macrophages in vivo via uptake by phagocytes was demonstrated. These results suggest that XL-MSNs are an effective platform for delivery of cytokines to MPS cells for in vivo modulation and the subsequent treatment of inflammation-associated diseases. Finally, anti-inflammatory nanoparticle assembly on the MSNs is developed. Cerium oxide (ceria) nanoparticles, inorganic ROS scavenging catalyst, synthesized in an aqueous media were electrostatically bound on the surface of MSNs that had magnetic nanoparticles for MR imaging contrast agents. In intracerebral hemorrhage (ICH) model, MRI clearly showed that the intracerebrally injected ceria nanoparticles doped MSNs was effectively delivered to the peri-hematomal area where secondary brain injury occurred. Furthermore, the lipid bilayer supported the multifunctional nanocomposition for increasing dispersity and circulation in body fluid. In the peri-hematomal area, multifunctional nanocomposition was internalized by macrophages and attenuated inflammatory cell infiltration through ROS scavenging capacity of ceria nanoparticles, which is required for resolving the intense inflammation induced after ICH. Furthermore, multifunctional nanocomposition significantly reduced brain edema induced by ICH and enhanced neurologic outcomes in treated rats. Taken together enhanced loading capacity and multifunctionalities of MSNs suggest that MSNs can be used as an attractive platform for immunotherapy in the future.

한국 생명의료기술의 전환에 관한 연구 : 재생산기술로부터 생명공학기술로

하정옥 서울대학교 대학원 2006 국내박사

Formation and Characterization of in vitro Bioengineered Neuromuscular Junction Models

Charoensook, Surapon N Columbia University ProQuest Dissertations & These 2018 해외박사(DDOD)

In vitro models of the neuromuscular junction (NMJ) are emerging as a valuable tool to study synaptogenesis, synaptic maintenance, and pathogenesis of neurodegenerative diseases. Many models have previously been developed using a variety of cell sources for muscle and motoneurons, but the models can be further improved by integrating beneficial features to better mimic the native milieu of NMJ development. We created a functional in vitro model of NMJ by bioreactor cultivation of C2C12 myoblasts, transdifferentiated myocytes and stem cell-derived motoneurons with electrical stimulation. Proper coculture medium and electrical stimulation led to improved functional coupling between the emerging motoneurons and myocytes, as evidenced by mature cellular structures, increased expression of neuronal and muscular genes, clusterization of acetylcholine receptors (AChRs) in the vicinity of motoneurons, and the response of the coculture to glutamate stimulation. To validate the models and demonstrate utility for pharmacological testing, we analyzed the potency of the drugs that affect key pathways during NMJ signal transduction, including acetylcholine (ACh) synthesis, ACh vesicular storage, ACh synaptic release, AChR activation, and ACh inactivation in the synaptic cleft. The models properly responded to the drugs in a concentration-dependent manner. The proposed in vitro NMJ model could thus be used in pharmacological screening and controlled studies of neuromuscular diseases.

Machine Learning Methods for Protein and Metabolic Engineering

Gupta, Sanjan T. P ProQuest Dissertations & Theses The University of 2020 해외박사(DDOD)

Recent advances in high-throughput technologies for probing biological systems coupled with increased interest in machine learning applications has led to a surge in data-driven bioengineering themed studies. This dissertation describes development and application of machine learning methods to solve data-scarce but yet combinatorially challenging protein and metabolic engineering problems. Additionally, these modeling problems are often characterized by very high-dimensional feature vector representations owing to that lack of good first principles based understanding of what makes one biomolecular design better than the other. To demonstrate proof-of-concept, these novel methods have been applied to various unsolved problems in the biomass to biofuels and chemicals pipeline.The first part of this dissertation describes MLProScape – a machine learning (ML) based method for engineering enzymes faster by modeling the protein fitness landscape (ProScape). Using MLProScape, one can build accurate models of protein fitness landscape to quantitatively predict the functional properties of an enzyme based on its protein sequence, and then, use them to identify a minimal set of sequence modifications (involving amino acid substitutions, insertions, and/or deletions) needed to design synthetic proteins with improved functional traits. MLProScape requires experimentally testing fewer protein variants (on the order of tens to hundreds) as against screening millions of protein variants in directed evolution approach and is capable of modeling complex design criteria, such as engineering the catalytic activity of an enzyme towards multiple substrates simultaneously. Moreover, the use of protein-wide position specific feature vector representation for modeling the protein sequences enables the identification of amino acid positions distal to the active site that might play a key role in modulating the activity level. As a proof-of-concept, MLProScape was applied to enhance the catalytic activity of glycoside hydrolases – an important enzyme used to degrade cellulosic and hemi-cellulosic components of lignocellulosic biomass. Using just one round of design-build-test cycle, several promising single point mutant designs were found that enhanced the cellulase and xylanase specific activities simultaneously.The next chapter describes the converse problem. By combining concepts from machine learning, cheminformatics, and enzyme kinetics, empirical models have been built to systematically exploit the substrate promiscuity of metabolic enzymes so as to eventually enable new biochemical reactions. Additionally, a novel optimization-based framework has been proposed for enabling meta-analysis. Using modeling gimmicks, the original MINLP (Mixed Integer Non-linear Programming) model was reformulated into simpler MIQCP (Mixed Integer Quadratically Constrained Programming) and MILP (Mixed Integer Linear Programming) formulations to aid computational tractability and convergence to optimality.The subsequent chapter further exploits the idea of promiscuity in the context of allosteric transcription factors (aTFs) to tweak the ligand specificity of transcription factors at an allosteric site as well as to design aTF-regulated de novo promoters. The pen-ultimate chapter describes ML statistical models as a means to understand and engineer mRNA transcripts with improved stability using biophysical properties and counts of sequence motifs in untranslated region as features, respectively. Insights and recommendations generated from these quantitative biology studies will in turn be beneficial for accelerating the bioengineering pipeline as well as improving the success rate for future rounds of biomolecular design.

Bioengineering Basics, Biometrology, and Biocivics with Ganoderma lucidum

Perez, Rolando Cruz ProQuest Dissertations & Theses Stanford Universit 2020 해외박사(DDOD)

Nanoscale bioengineered biomolecules for ultrasensitive optical biosensors

Chavan, Sachin Ganpat 중앙대학교 대학원 2021 국내박사

초민감 광학 바이오센서를 위한 나노스케일 바이오엔지니어링 바이오 분자

샤힌 샤반 중앙대학교 대학원 2021 국내박사

An optical sensor is currently receiving tremendous attention in a bio-medical technology to required time-intensive and specific quantification biosensor demanding for reliable, inexpensive, and highly sensitive devices at the point of care. Thus, a versatile, nanosized, genetically bioengineered human apoferritin heavy chain (hAFTN-H) protein scaffold nanoparticle was synthesized for biosensor applications. In this study, hAFTN-H consisting of 24 subunits, was genetically expressed with protein-G and 6x His-tag adaptor units. The gene was incorporated into pET-28b(+) plasmid at the C-terminal site of Escherichia Coli (E. Coli). This nanoconstruct scaffold is resulting in the biosensing strategy based on the optical platform. We proposed a straightforward and label-free, highly sensitive biosensor to quantifying Infectious Pancreatic Necrosis Virus (IPNV) based on hAFTN-H nanoparticles for analyte detection by fluorometry colorimetry (lateral flow) and colorimetric switching methods. In a fluorometry detection, we achieved a two-step signal amplifying strategy that employed a recombinant hAFTN nanoparticle modified to express a protein G bound to the targeted Fc region of monoclonal antibodies and a 6×His-tag that vigorously collaborates with the functionalized Ni-NTA moieties. The magnetic beads used for rapid separation chemically tactic attempted to target IPNV fish virus via anti-capture and detection IPNV antibodies. The arrangement of hAFTN-H nanoprobe surface-expressed subunits allows binding anti-detection monoclonal antibody and Ni-NTA Atto fluorescent dye via protein G 6xHis-tag derivatives. Further, the experiment of lateral flow-based colorimetry assay offered a new opportunity for readily distinguishable quantification of IPNV involving due to the high sensitivity of detection demands. Herein hAFTN-H nanoprobe outer surface generated cross-linkage with 5 nm Ni-NTA gold nanoparticle complex efficiently monitoring to visualization and quantification towards the IPNV system. The proposed fluorimetry system exhibit detection limits towards IPNV 1.02 TCID50/mL (50% tissue culture infective dose), and colorimetric (lateral flow) was 0.88 In our present study, we were promoting a very tiny biomolecule enzyme-mediated physiological reaction between neurotransmitter serotonin (5-HT) oxidation to the formation of a dimer complex. 5-HT acceleration of catalytic activity to formation dimer at C-4 position to the formed phenolic radical presence of peroxidase and H2O2. The dimer 5-HT possesses the capability to recognize intermolecular interaction units that cause aggregation complex of Au nanoparticles (Au-NPs) for logically inexpensive and straightforward analytical demands. As proof of visual and spectral analysis, peroxidative dimeric 5-HT demonstrated a sensitive, robust, low-cost, and sensible immunosensor, represents the first-time quantifying at point of care (POC) platform. The calorimetric method enables highly sensitive detection of 5-HT in phosphate buffer samples at nanomolar levels with an LOD of 2.6 nM, and the sensor possesses a dynamic range of 100-300 nM in buffer conditions. 광학 센서는 현장진단 분야에서 신뢰할 수 있고 저렴하며 매우 민감한 장비를 필요로 하는 바이오메디컬 기술에 있어서 현재 많은 주목을 받고 있다. 따라서, 바이오센서로 사용하기 위해 다용도 나노 사이즈의 유전적으로 변형된 인간 아포페리틴 중사슬 (hAFTN-H) 단백질 스캐폴드 나노입자가 합성되었다. 본 연구에서는 24개의 하위단위로 구성된 hAFTN-H는 단백질-G와 6x His-tag 어댑터 단위와 결합되어 유전적으로 발현되었다. hAFTN-H의 유전자는 대장균(E. Coli)의 C-말단부위에서 pET-28b(+) 플라스미드에 삽입되었다. 이 나노구조 스캐폴드는 광학 플랫폼에 기반한 바이오센싱 전략을 만들어내고 있다. 형광, 흡광 (측방 유동 측정) 및 색 변환 측정법에 의한 타겟 측정을 위해서 hAFTN-H 나노입자를 기반으로 전염성 췌장 괴사 바이러스 (IPNV)를 정량화하기 위한 간단하면서도 라벨이 없는 매우 민감한 바이오센서를 제안했다. 본 연구에서는 단일 클론 항체의 표적 Fc영역에 결합된 단백질 G와 Ni-NTA 부분과 강력하게 결합하는 6x His-tag를 발현을 확인하기 위한 형광측정에서 수정된 재조합 hAFTN 나노입자를 이용하여 2단계 신호 증폭을 확인했다. 신속한 분리를 위해 사용된 자성입자는 IPNV 포획 및 검출 항체와 결합되어 IPNV 어류 바이러스를 잡기위해 적용되었다. hAFTN-H 나노프로브 표면에 발현된 하위단위의 배열은 단백질 G와 6x His-tag 유도체를 통해 항-단일 검출항체와 Ni-NTA Atto 형광 염료를 결합하게 한다. 또한, 측방 유동에 기반한 흡광 측정 실험은 높은 측정 민감도로 IPNV를 쉽게 구별가능한 정량화를 위한 새로운 기회를 제공하였다. 여기에서는 hAFTN-H 나노프로브 외부 표면이 IPNV 시스템에 대한 시각화 및 정량화를 효율적으로 모니터링하는 5nm 크기의 Ni-NTA 금 나노입자 복합체와의 가교결합을 발생시켰다. 제안된 형광 시스템은 IPNV에 대하여 1.02 TCID50/mL (50% 조직배양 감염 선량)의 검출 한계를 보였고 흡광 (측방 유동) 측정은 0.88의 검출한계를 보였다. 본 연구에서, 다이머 복합체의 형성으로 신경전달물질 세로토닌 (5-HT) 산화 사이의 매우 작은 생체분자 효소 매개 생리학적 반응을 촉진하였다. C-4 위치에서 다이머 형성에 대한 촉매반응의 5-HT 가속화는 페놀릭 라디칼 존재인 과산화효소 및 과산화수소를 형성했다. 다이머 5-HT는 논리적으로 저렴하고 간단한 분석 수요에 대해 금 나노입자 (Au-NPs)의 응집 복합체를 유발하는 분자간 상호작용 단위를 인식할 수 있는 능력을 가지고 있다. 시각적 및 스펙트럼 분석의 증거로서 과산화 다이머 5-HT는 민감하고 강력하며 저비용이며 합리적인 면역센서를 입증했으며 현장진단 플랫폼에서 최초의 정량화를 보였다. 흡광 측정법은 인산염 완충용액에 있는 5-HT를 2.6 nM의 검출한계를 가진 나노몰 단위 수준의 매우 민감한 측정을 가능하게 했고 완충용액 조건에서 100 – 300 nM의 측정 범위를 가지고 있다.