BioTechniques最新文献

Dual compartment utility of BRET-based biosensors for PPP2R5A/B56α, a cancer-associated B regulatory subunit of PP2A. 基于bret的双室生物传感器PPP2R5A/B56α， PP2A的癌症相关B调节亚基。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-06-27 DOI: 10.1080/07366205.2025.2523093

Hirofumi Yamauchi, Atsuro Oishi, Masahiko Ajiro, Atsuhito Nakayama, Kazuki Nishimura, Michiko Kurikawa, Mina Yoshida, Rei Kudo, Minori Koizumi, Takuya Izumi-Tamura, Miki Nagase, Natsuko Shinohara, Mayumi Hanzawa, Marimu Sakumoto, Takahiro Nishino, Ryoichi Maenosono, Asuka Kawachi, Junko Mukohyama, Shingo Yano, Tomoya Muto, Akihide Yoshimi

{"title":"Dual compartment utility of BRET-based biosensors for PPP2R5A/B56α, a cancer-associated B regulatory subunit of PP2A.","authors":"Hirofumi Yamauchi, Atsuro Oishi, Masahiko Ajiro, Atsuhito Nakayama, Kazuki Nishimura, Michiko Kurikawa, Mina Yoshida, Rei Kudo, Minori Koizumi, Takuya Izumi-Tamura, Miki Nagase, Natsuko Shinohara, Mayumi Hanzawa, Marimu Sakumoto, Takahiro Nishino, Ryoichi Maenosono, Asuka Kawachi, Junko Mukohyama, Shingo Yano, Tomoya Muto, Akihide Yoshimi","doi":"10.1080/07366205.2025.2523093","DOIUrl":"https://doi.org/10.1080/07366205.2025.2523093","url":null,"abstract":"Protein phosphatase 2A (PP2A), a pivotal serine/threonine phosphatase, plays a crucial role in cellular regulation and tumor suppression. Dysregulation of PP2A complex, particularly the Aα subunit and B56 family, is linked to malignancies through altered substrate interactions, exemplified by c-MYC dynamics. Given the challenges in identifying PP2A substrates-owing to the enzyme's expansive substrate range, transient interaction profiles, and complex regulatory mechanisms-we employed bioluminescence resonance energy transfer (BRET) sensors. These advanced molecular tools facilitate the real-time detection of protein-protein interactions within live cells. This investigation details the creation and application of a novel PPP2R5A (B56α) BRET sensor tailored for cytosolic and nuclear environments, effectively distinguishing specific PP2A interactions. The nuclear sensor, enhanced with a nuclear localization signal, enabled probing of targets like c-MYC. The dual compartmental utility of these sensors underscores their significant potential in elucidating PP2A's regulatory roles and their implications in oncogenesis. Our study highlights the efficacy of BRET sensors in formulating precision therapeutic strategies. This advancement provides a robust framework for deeper investigations into the multifaceted roles of PP2A in both normal physiological and pathological contexts, paving the way for future explorations into its intricate molecular interactions.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"1-11"},"PeriodicalIF":2.2,"publicationDate":"2025-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144526396","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Mito-Kaede photoactivation and chase experiment for mitophagy: mitophagy flux response toward various stimulations. Mito-Kaede对线粒体自噬的光激活与追逐实验：线粒体自噬通量对各种刺激的响应。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-05-31 DOI: 10.1080/07366205.2025.2505357

Yoh Sugawara, Hiroyuki Morinaga, Jingyuan Chen, Yoshinori Kitagawa, Hiroki Ogata, Asiya Karim, Miu Kikuchi, Maryam Khan, Erica Yasuhara, Takahisa Goto, Joseph A Jeevendra Martyn, Shingo Yasuhara

引用次数: 0

Integration of mass cytometry and single-cell RNA-sequencing of cells in bronchoalveolar lavage. 支气管肺泡灌洗细胞的细胞细胞计数和单细胞rna测序的整合。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-05-19 DOI: 10.1080/07366205.2025.2505347

Elizabeth Guinto, Sameeksha Chopra, I-Chih Kuo, Samuel B Shin, Melina Messing, Chung Y Cheung, Julia Sw Yang, Firoozeh V Gerayeli, William Yip, Stephen Milne, Rachel L Eddy, Janice M Leung, Kelly M McNagny, Don D Sin

引用次数: 0

Correction. 修正。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-01-16 DOI: 10.1080/07366205.2025.2450187

引用次数: 0

Freezing diluted bovine serum albumin standards does not significantly affect standard curves. 冷冻稀释牛血清白蛋白标准品对标准曲线影响不显著。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-05-23 DOI: 10.1080/07366205.2025.2502268

Rachelle Sheets, Bhavik Rajaboina, Caitlin E Bromberg, Liam P Curtin, Mitchell L Haddock, Phillip Stafford, Theresa Currier Thomas, Adrienne C Scheck

{"title":"Freezing diluted bovine serum albumin standards does not significantly affect standard curves.","authors":"Rachelle Sheets, Bhavik Rajaboina, Caitlin E Bromberg, Liam P Curtin, Mitchell L Haddock, Phillip Stafford, Theresa Currier Thomas, Adrienne C Scheck","doi":"10.1080/07366205.2025.2502268","DOIUrl":"10.1080/07366205.2025.2502268","url":null,"abstract":"Total protein isolation followed by quantitation using a colorimetric method, such as the bicinchoninic acid (BCA) assay is a common laboratory protocol. Protein concentrations are determined by comparing extracted samples to a standard curve generated from serial dilutions of a reference protein, such as bovine serum albumin (BSA). This study aimed to identify the most reproducible and accurate method for quantifying protein concentrations in an experimental series over time. We analyzed the effect of serial freeze-thaws, inter-person and intra-person variability in standard preparation and assay execution. Absorbance was measured at 565 nanometers (nm) using an Epoch Microplate Spectrophotometer (Agilent Technologies, Inc., Santa Clara, CA) with Gen5 Data Analysis software. The most consistent and accurate method for determining the protein concentrations over time is to prepare a large batch of diluted BSA standards, aliquot them into small portions, and store them frozen.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"77 3","pages":"95-102"},"PeriodicalIF":2.2,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12222612/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144126559","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Standardized droplet preamplification method for downstream circulating cell-free DNA analysis. 标准化滴滴预扩增法用于下游循环无细胞DNA分析。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-05-23 DOI: 10.1080/07366205.2025.2504287

Colin Skeen, Erica D Pratt

{"title":"Standardized droplet preamplification method for downstream circulating cell-free DNA analysis.","authors":"Colin Skeen, Erica D Pratt","doi":"10.1080/07366205.2025.2504287","DOIUrl":"https://doi.org/10.1080/07366205.2025.2504287","url":null,"abstract":"Circulating cell-free DNA (ccfDNA) can be found in blood and other biofluids and is a minimally invasive biomarker for several pathological processes. As tumors become more invasive, an increasing amount of circulating tumor DNA (ctDNA) is also shed into the peripheral circulation. Combined analysis of ccfDNA and ctDNA has demonstrated prognostic and predictive value in metastatic disease. However, localized tumors shed significantly less ccfDNA/ctDNA and accurate detection remains a technical challenge. To overcome this barrier, droplet preamplification has been used to perform robust multiplexed analysis of low-input samples. To reduce false positives, it is essential to use a high-fidelity polymerase with 3'-5' exonuclease activity. However, attempts to combine high-fidelity polymerases with commercial droplet digital chemistries have had limited success. There is also no standardized method for efficient amplicon recovery from droplets. In this work, we present a method to reliably stabilize emulsions and recover preamplified templates. We systematically compared our protocol with different destabilization methods and found an average 41% improvement in recovery efficiency. We anticipate that this standardized method will increase the consistency and reproducibility of ccfDNA/ctDNA analyses. This technique could be readily translated to other low-input or low-biomass samples, such as urine, saliva, or archived biopsy specimens.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":"77 3","pages":"125-135"},"PeriodicalIF":2.2,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144126564","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Developing an insert to protect large DNA molecules during cell lysis. 开发一种插入物，在细胞裂解过程中保护大的DNA分子。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-03-26 DOI: 10.1080/07366205.2025.2483601

Charles Polen, Esmeralda Mendez Ortiz, Cordelia Harbison, Nate Garringer, Martin Lopez, Kristy L Kounovsky-Shafer

引用次数: 0

Efficient spheroid morphology assessment with a ChatGPT data analyst: implications for cell therapy. 高效球体形态评估与ChatGPT数据分析：对细胞治疗的影响。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-04-23 DOI: 10.1080/07366205.2025.2493489

Takuya Sakamoto, Hiroto Koma, Ayane Kuwano, Tetsuhiro Horie, Atsushi Fuku, Hironori Kitajima, Yuka Nakamura, Ikuhiro Tanida, Yujiro Nakade, Hiroaki Hirata, Yoshiyuki Tachi, Hiroshi Sunami, Daisuke Sakamoto, Sohsuke Yamada, Naoki Yamamoto, Yusuke Shimizu, Yasuhito Ishigaki, Toru Ichiseki, Ayumi Kaneuji, Satoshi Osawa, Norio Kawahara

{"title":"Efficient spheroid morphology assessment with a ChatGPT data analyst: implications for cell therapy.","authors":"Takuya Sakamoto, Hiroto Koma, Ayane Kuwano, Tetsuhiro Horie, Atsushi Fuku, Hironori Kitajima, Yuka Nakamura, Ikuhiro Tanida, Yujiro Nakade, Hiroaki Hirata, Yoshiyuki Tachi, Hiroshi Sunami, Daisuke Sakamoto, Sohsuke Yamada, Naoki Yamamoto, Yusuke Shimizu, Yasuhito Ishigaki, Toru Ichiseki, Ayumi Kaneuji, Satoshi Osawa, Norio Kawahara","doi":"10.1080/07366205.2025.2493489","DOIUrl":"10.1080/07366205.2025.2493489","url":null,"abstract":"Background: Adipose-derived stem cells (ADSCs) exhibit promising potential for the treatment of various diseases, including osteoarthritis. Spheroids derived from ADSCs are a viable treatment option with enhanced anti-inflammatory effects and tissue repair capabilities.Objective: SphereRing® is a rotating donut-shaped tube that efficiently produces large quantities of spheroids. However, accurately measuring spheroid size for spheroid quality assessment is challenging. This study aimed to develop an automated method for measuring spheroid size using deep learning through the ChatGPT Data Analyst for image recognition and processing.Method: The area, perimeter, and circularity of spheroids generated with the SphereRing system were analyzed using ChatGPT Data Analyst and ImageJ. Measurement accuracy was validated using Bland-Altman analysis and scatter plot correlation coefficients.Results: ChatGPT Data Analyst was consistent with ImageJ for all parameters. Bland-Altman plots demonstrated strong agreement; most data points were within the 95% limits.Conclusion: The ChatGPT Data Analyst provides a reliable and efficient alternative for assessing spheroid quality. This method reduces human error and improves reproducibility to enhance spheroid quality control. Thus, this method has potential applications in regenerative medicine.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"137-149"},"PeriodicalIF":2.2,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143958147","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A drug screening platform for protein expression levels in neurological disorders. 神经系统疾病蛋白表达水平的药物筛选平台。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-03-01 Epub Date: 2025-04-03 DOI: 10.1080/07366205.2025.2484094

Farida Emran, Ibrahim Kays, Chiu-An Lo, Yueyang Li, Brian E Chen

引用次数: 0

Impact of commercial RNA extraction methods on the recovery of human RNA sequence data from archival fixed tissues. 商业RNA提取方法对从档案固定组织中恢复人类RNA序列数据的影响。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-02-01 Epub Date: 2025-03-12 DOI: 10.1080/07366205.2025.2473842

Tess Wilson, Melanie Kuch, Debi Poinar, Jasmine Rockarts, Bruce Wainman, Susan Morgello, Hendrik Poinar

{"title":"Impact of commercial RNA extraction methods on the recovery of human RNA sequence data from archival fixed tissues.","authors":"Tess Wilson, Melanie Kuch, Debi Poinar, Jasmine Rockarts, Bruce Wainman, Susan Morgello, Hendrik Poinar","doi":"10.1080/07366205.2025.2473842","DOIUrl":"10.1080/07366205.2025.2473842","url":null,"abstract":"Archival fixed tissues hold key insights into the evolutionary history of RNA viruses and the associated host immune response, yet access to the RNA sequence data is limited by a lack of robust methods for RNA extraction and sequence retrieval from these tissue types. Here we compared three commercial RNA extraction techniques (bead, column, and phase-based) on five fixed human brain tissues done in triplicate, that have been stored for up to 43 years. We found that for this sample set, bead-based extractions captured longer molecules and yielded a greater proportion of unique reads when aligned to the human genome, than did column and phase-based extraction methods. Via the incorporation of multiple extraction replicates, we quantified the variability in sequencing metrics resulting from tissue sample and extraction technique heterogeneity. Additionally, we compared pre- and post-sequencing metrics and found that the former poorly predicted post-sequencing on-target success. Our findings help inform future research on the recovery of RNA from archival fixed tissues.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"76-93"},"PeriodicalIF":2.2,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12063700/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143603124","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0