Biology Methods and Protocols最新文献_第9页

An optimized protocol for generating appendage-bearing skin organoids from human-induced pluripotent stem cells. 利用人体诱导多能干细胞生成附肢皮肤器官组织的优化方案。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-03-22 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae019

Imaan Ahmed, Jane Sun, Jason Brown, Kiarash Khosrotehrani, Abbas Shafiee

引用次数: 0

Alternative community-led intervention to improve uptake of cataract surgery services in rural Tanzania-The Dodoma Community Cataract Acceptance Trial (DoCCAT): a protocol for intervention co-designing and implementation in a cluster-randomized controlled trial. 提高坦桑尼亚农村地区白内障手术服务接受率的替代性社区主导干预措施--多多马社区白内障接受试验（DoCCAT）：群组随机对照试验中干预措施的共同设计和实施方案。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-03-08 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae016

Frank Sandi, Gareth Mercer, Robert Geneau, Kenneth Bassett, Deogratius Bintabara, Albino Kalolo

{"title":"Alternative community-led intervention to improve uptake of cataract surgery services in rural Tanzania-The Dodoma Community Cataract Acceptance Trial (DoCCAT): a protocol for intervention co-designing and implementation in a cluster-randomized controlled trial.","authors":"Frank Sandi, Gareth Mercer, Robert Geneau, Kenneth Bassett, Deogratius Bintabara, Albino Kalolo","doi":"10.1093/biomethods/bpae016","DOIUrl":"https://doi.org/10.1093/biomethods/bpae016","url":null,"abstract":"Age-related lens opacification (cataract) remains the leading cause of visual impairment and blindness worldwide. In low- and middle-income countries, utilization of cataract surgical services is often limited despite community-based outreach programmes. Community-led research, whereby researchers and community members collaboratively co-design intervention is an approach that ensures the interventions are locally relevant and that their implementation is feasible and socially accepted in the targeted contexts. Community-led interventions have the potential to increase cataract surgery uptake if done appropriately. In this study, once the intervention is co-designed it will be implemented through a cluster-randomized controlled trial (cRCT) with ward as a unit of randomization. This study will utilise both the qualitative methods for co-designing the intervention and the quantitative methods for effective assessment of the developed community-led intervention through a cRCT in 80 rural wards of Dodoma region, Tanzania (40 Intervention). The 'intervention package' will be developed through participatory community meetings and ongoing evaluation and modification of the intervention based on its impact on service utilization. Leask's four stages of intervention co-creation will guide the development within Rifkin's CHOICE framework. The primary outcomes are two: the number of patients attending eye disease screening camps, and the number of patients accepting cataract surgery. NVivo version 12 will be used for qualitative data analysis and Stata version 12 for quantitative data. Independent and paired t-tests will be performed to make comparisons between and within groups. P-values less than 0.05 will be considered statistically significant.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae016"},"PeriodicalIF":3.6,"publicationDate":"2024-03-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10987207/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140865230","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A rapid and accurate method for evaluating the degradation of pan-Akt in cells by PROTACs using NanoLuc luciferase. 利用 NanoLuc 荧光素酶快速准确地评估 PROTACs 在细胞中降解 pan-Akt 的方法。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-03-05 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae014

Xiaojun Ji, Lei Miao, Yebin Wu, Tingli Zhao, Yaxuan Si, Xiaoyun Tan, Qiuhua Zhou, Rui Zuo, Junjie Pei, Jian Wu, Changyou Ma, Zhongjun Ma, Dan Xu

{"title":"A rapid and accurate method for evaluating the degradation of pan-Akt in cells by PROTACs using NanoLuc luciferase.","authors":"Xiaojun Ji, Lei Miao, Yebin Wu, Tingli Zhao, Yaxuan Si, Xiaoyun Tan, Qiuhua Zhou, Rui Zuo, Junjie Pei, Jian Wu, Changyou Ma, Zhongjun Ma, Dan Xu","doi":"10.1093/biomethods/bpae014","DOIUrl":"10.1093/biomethods/bpae014","url":null,"abstract":"Proteolysis targeting chimera (PROTAC) is a protein degradation technique that has been increasingly used in the development of new drugs in recent years. Akt is a classical serine/threonine kinase, and its role outside of the kinase has gradually gained attention in recent years, making it one of the proteins targeted by PROTACs. Currently, there are many methods used for the evaluation of intracellular protein degradation, but each has its own advantages or disadvantages. This study aimed to investigate the feasibility of evaluating the degradation of pan-Akt proteins in cells by PROTACs (MS21 and MS170) using the NanoLuc luciferase method. After conducting a thorough comparison between this method and the classical western blot assay in various cells, as well as testing the stability of the experiments between multiple batches, we found that NanoLuc luciferase is a highly accurate, stable, low-cost and easy-to-operate method for the evaluation of intracellular pan-Akt degradation by PROTACs with a short cycle time and high cellular expandability. Given the numerous advantages of this method, it is hypothesized that it could be extended to evaluate the degradation of more target proteins of PROTACs. In summary, the NanoLuc luciferase is a suitable method for early protein degradation screening of PROTAC compounds.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae014"},"PeriodicalIF":3.6,"publicationDate":"2024-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10965420/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140307242","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

An improved method for measuring catalase activity in biological samples. 测量生物样本中过氧化氢酶活性的改进方法。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-03-05 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae015

Mahmoud Hussein Hadwan, Marwah Jaber Hussein, Rawa M Mohammed, Asad M Hadwan, Hawraa Saad Al-Kawaz, Saba S M Al-Obaidy, Zainab Abbas Al Talebi

{"title":"An improved method for measuring catalase activity in biological samples.","authors":"Mahmoud Hussein Hadwan, Marwah Jaber Hussein, Rawa M Mohammed, Asad M Hadwan, Hawraa Saad Al-Kawaz, Saba S M Al-Obaidy, Zainab Abbas Al Talebi","doi":"10.1093/biomethods/bpae015","DOIUrl":"10.1093/biomethods/bpae015","url":null,"abstract":"Catalase (CAT) is an important enzyme that protects biomolecules against oxidative damage by breaking down hydrogen peroxide (H2O2) into water and oxygen. CAT is present in all aerobic microbes, animals, and plants. It is, however, absent from normal human urine but can be detected in pathological urine. CAT testing can thus help to detect such urine. This study presents a novel spectrophotometric method for determining CAT activity characterized by its simplicity, sensitivity, specificity, and rapidity. The method involves incubating enzyme-containing samples with a carefully chosen concentration of H2O2 for a specified incubation period. Subsequently, a solution containing ferrous ammonium sulfate (FAS) and sulfosalicylic acid (SSA) is added to terminate the enzyme activity. A distinctive maroon-colored ferrisulfosalicylate complex is formed. The formation of this complex is a direct result of the reaction between FAS and any residual peroxide present. This leads to the generation of ferric ions when coordinated with SSA. The complex has a maximum absorbance of 490 nm. This advanced method eliminates the need for concentrated acids to stop CAT activity, making it safer and easier to handle. A comparative analysis against the standard ferrithiocyanate method showed a correlation coefficient of 0.99, demonstrating the new method's comparable effectiveness and reliability. In conclusion, a simple and reliable protocol for assessing CAT activity, which utilizes a cuvette or microplate, has been demonstrated in this study. This interference-free protocol can easily be used in research and clinical analysis with considerable accuracy and precision.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae015"},"PeriodicalIF":3.6,"publicationDate":"2024-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10957919/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140207689","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Pairing a bioinformatics-focused course-based undergraduate research experience with specifications grading in an introductory biology classroom. 在生物学入门课堂上，将以生物信息学为重点的课程为基础的本科生研究体验与规范评分相结合。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-02-28 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae013

Melinda A Yang, Kylie Korsnack

{"title":"Pairing a bioinformatics-focused course-based undergraduate research experience with specifications grading in an introductory biology classroom.","authors":"Melinda A Yang, Kylie Korsnack","doi":"10.1093/biomethods/bpae013","DOIUrl":"10.1093/biomethods/bpae013","url":null,"abstract":"Introducing bioinformatics-focused concepts and skills in a biology classroom is difficult, especially in introductory biology classrooms. Course-based Undergraduate Research Experiences (CUREs) facilitate this process, introducing genomics and bioinformatics through authentic research experiences, but the many learning objectives needed in scientific research and communication, foundational biology concepts, and bioinformatics-focused concepts and skills can make the process challenging. Here, the pairing of specifications grading with a bioinformatics-focused CURE developed by the Genomics Education Partnership is described. The study examines how the course structure with specifications grading facilitated scaffolding of writing assignments, group work, and metacognitive activities; and describes the synergies between CUREs and specifications grading. CUREs require mastery of related concepts and skills for working through the research process, utilize common research practices of revision and iteration, and encourage a growth mindset to learning-all of which are heavily incentivized in assessment practices focused on specifications grading.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae013"},"PeriodicalIF":3.6,"publicationDate":"2024-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10924719/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140094809","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Correction to: Long amplicon nanopore sequencing of Botrytis cinerea and other fungal species present in infected grapevine leaf samples. 更正：长扩增片段纳米孔测序受感染葡萄叶片样本中的灰葡萄孢和其他真菌物种。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-02-23 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae010

引用次数: 0

Rapid IDH1-R132 genotyping panel utilizing locked nucleic acid loop-mediated isothermal amplification. 利用锁定核酸环介导等温扩增技术快速进行 IDH1-R132 基因分型检测。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-02-21 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae012

Kristian A Choate, Edward J Raack, Paul B Mann, Evan A Jones, Robert J Winn, Matthew J Jennings

{"title":"Rapid IDH1-R132 genotyping panel utilizing locked nucleic acid loop-mediated isothermal amplification.","authors":"Kristian A Choate, Edward J Raack, Paul B Mann, Evan A Jones, Robert J Winn, Matthew J Jennings","doi":"10.1093/biomethods/bpae012","DOIUrl":"https://doi.org/10.1093/biomethods/bpae012","url":null,"abstract":"While the detection of single-nucleotide variants (SNVs) is important for evaluating human health and disease, most genotyping methods require a nucleic acid extraction step and lengthy analytical times. Here, we present a protocol which utilizes the integration of locked nucleic acids (LNAs) into self-annealing loop primers for the allelic discrimination of five isocitrate dehydrogenase 1 R132 (IDH1-R132) variants using loop-mediated isothermal amplification (LAMP). This genotyping panel was initially evaluated using purified synthetic DNA to show proof of specific SNV discrimination. Additional evaluation using glioma tumor lysates with known IDH1-R132 mutational status demonstrated specificity in approximately 35 min without the need for a nucleic acid extraction purification step. This LNA-LAMP-based genotyping assay can detect single base differences in purified nucleic acids or tissue homogenates, including instances where the variant of interest is present in an excess of background wild-type DNA. The pH-based colorimetric indicator of LNA-LAMP facilitates convenient visual interpretation of reactions, and we demonstrate successful translation to an end-point format using absorbance ratio, allowing for an alternative and objective approach for differentiating between positive and negative reactions. Importantly, the LNA-LAMP genotyping panel is highly reproducible, with no false-positive or false-negative results observed.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae012"},"PeriodicalIF":3.6,"publicationDate":"2024-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10984729/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140871390","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Development of a highly sensitive TaqMan method based on multi-probe strategy: its application in ASFV detection. 基于多探针策略的高灵敏度 TaqMan 方法的开发：在 ASFV 检测中的应用。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-02-19 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae011

Shuxiang Ding, Tianren Shen, Zixuan Feng, Sujing Diao, Yan Yan, Zhenkun Du, Yulan Jin, Jinyan Gu, Jiyong Zhou, Min Liao, Weiren Dong

{"title":"Development of a highly sensitive TaqMan method based on multi-probe strategy: its application in ASFV detection.","authors":"Shuxiang Ding, Tianren Shen, Zixuan Feng, Sujing Diao, Yan Yan, Zhenkun Du, Yulan Jin, Jinyan Gu, Jiyong Zhou, Min Liao, Weiren Dong","doi":"10.1093/biomethods/bpae011","DOIUrl":"10.1093/biomethods/bpae011","url":null,"abstract":"The establishment of high sensitive detection method for various pathogenic microorganisms remains constantly concerned. In the present study, multi-probe strategy was first systematically investigated followed by establishing a highly sensitive TaqMan real-time fluorescent quantitative PCR (qPCR) method for detecting African swine fever virus (ASFV). Briefly, four probes based on the B646L gene of ASFV were designed and the effects of different combinations of the probes in a single TaqMan qPCR assay on the detection sensitivity were investigated. As less as 0.5-5 copies/μl of the ASFV gene was detected by the established TaqMan qPCR assay. Furthermore, plasmid harboring the B646L in water samples could be concentrated 1000 times by ultrafiltration to enable a highly sensitive detection of trace viral nucleic acids. Moreover, no cross-reactivity was observed with other common clinical swine viruses such as PCV2, PCV3, PCV4, PEDV, PDCoV, CSFV, PRRSV, and PRV. When detecting 173 clinical porcine serum samples, the coincidence rate between the developed method and WOAH (World Organization of Animal Health) recommended method was 100%. This study might provide an integrated strategy to achieve higher detection sensitivity of trace pathogenic microorganisms and applicably sensitive TaqMan-based qPCR assays.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae011"},"PeriodicalIF":3.6,"publicationDate":"2024-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10939455/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140132771","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A methodological primer of extracellular vesicles isolation and characterization via different techniques. 通过不同技术分离和表征细胞外囊泡的方法论入门。

IF 3.6

Biology Methods and Protocols Pub Date : 2024-02-13 eCollection Date: 2024-01-01 DOI: 10.1093/biomethods/bpae009

Farhang Aliakbari, Noah B Stocek, Maxximuss Cole-André, Janice Gomes, Giovanni Fanchini, Stephen H Pasternak, Gunna Christiansen, Dina Morshedi, Kathryn Volkening, Michael J Strong

{"title":"A methodological primer of extracellular vesicles isolation and characterization via different techniques.","authors":"Farhang Aliakbari, Noah B Stocek, Maxximuss Cole-André, Janice Gomes, Giovanni Fanchini, Stephen H Pasternak, Gunna Christiansen, Dina Morshedi, Kathryn Volkening, Michael J Strong","doi":"10.1093/biomethods/bpae009","DOIUrl":"10.1093/biomethods/bpae009","url":null,"abstract":"We present four different protocols of varying complexity for the isolation of cell culture-derived extracellular vesicles (EVs)/exosome-enriched fractions with the objective of providing researchers with easily conducted methods that can be adapted for many different uses in various laboratory settings and locations. These protocols are primarily based on polymer precipitation, filtration and/or ultracentrifugation, as well as size-exclusion chromatography (SEC) and include: (i) polyethylene glycol and sodium chloride supplementation of the conditioned medium followed by low-speed centrifugation; (ii) ultracentrifugation of conditioned medium; (iii) filtration of conditioned media through a 100-kDa exclusion filter; and (iv) isolation using a standard commercial kit. These techniques can be followed by further purification by ultracentrifugation, sucrose density gradient centrifugation, or SEC if needed and the equipment is available. HEK293 and SH-SY5Y cell cultures were used to generate conditioned medium containing exosomes. This medium was then depleted of cells and debris, filtered through a 0.2-µM filter, and supplemented with protease and RNAse inhibitors prior to exosomal isolation. The purified EVs can be used immediately or stably stored at 4°C (up to a week for imaging or using intact EVS downstream) or at -80°C for extended periods and then used for biochemical study. Our aim is not to compare these methodologies but to present them with descriptors so that researchers can choose the \"best method\" for their work under their individual conditions.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"9 1","pages":"bpae009"},"PeriodicalIF":3.6,"publicationDate":"2024-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10902684/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139997669","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Teaching at the intersection of science and society: an activity on healthcare disparities 科学与社会交汇处的教学：关于医疗保健差异的活动

IF 3.6

Biology Methods and Protocols Pub Date : 2024-01-05 DOI: 10.1093/biomethods/bpad041

Paula E Adams, Enya Granados, Abby E. Beatty, C. Ballen

{"title":"Teaching at the intersection of science and society: an activity on healthcare disparities","authors":"Paula E Adams, Enya Granados, Abby E. Beatty, C. Ballen","doi":"10.1093/biomethods/bpad041","DOIUrl":"https://doi.org/10.1093/biomethods/bpad041","url":null,"abstract":"\u0000 Understanding the relationship between science and society is an objective of science education and included as a core competency in the AAAS Vision and Change guidelines for biology education. However, traditional undergraduate biology instruction emphasizes scientific practice and generally avoids potentially controversial issues at the intersection of biology and society. By including these topics in biology coursework, instructors can challenge damaging ideologies and systemic inequalities that have influenced science, such as biological essentialism and health disparities. Specifically, an ideologically aware curriculum highlights how ideologies and paradigms shape our biological knowledge base and the application of that knowledge. Ideologically aware lessons emphasize the relationship between science and society with an aim to create more transparent, scientifically accurate, and inclusive postsecondary biology classrooms. Here we expand upon our ideologically aware curriculum with a new activity that challenges undergraduate biology students to consider the impacts of healthcare disparities. This lesson allows instructors to directly address systemic inequalities and allows students to connect biomedical sciences to real-world issues. Implementing an ideologically aware curriculum enables students to challenge prevailing worldviews and better address societal problems that lead to exclusion and oppression.","PeriodicalId":36528,"journal":{"name":"Biology Methods and Protocols","volume":"24 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2024-01-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139383809","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0