BioTechniques最新文献

Parameter optimization of NanoJ-SRRF for live-cell microtubule imaging. NanoJ-SRRF活细胞微管成像参数优化。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-07-22 DOI: 10.1080/07366205.2025.2534301

Sanhua Fang, Li Liu, Dan Yang, Shuangshuang Liu, Junli Xuan, Guifeng Xiao, Qianbing Zhao

引用次数: 0

A novel device for buffy coat collection. 一种用于收集灰褐色大衣的新装置。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-07-22 DOI: 10.1080/07366205.2025.2534299

Kyle Christian Hardy, Lisa Arrigo, Jennifer Campbell, Jacob Fiedler, Paul Fiedler

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 : 2025-04-01 Epub 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

Assessing the necessity of technical replicates in reverse transcription quantitative PCR. 评价反转录定量PCR中技术重复的必要性。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-04-01 Epub Date: 2025-07-05 DOI: 10.1080/07366205.2025.2527536

Eleni Christoforidou, Majid Hafezparast

{"title":"Assessing the necessity of technical replicates in reverse transcription quantitative PCR.","authors":"Eleni Christoforidou, Majid Hafezparast","doi":"10.1080/07366205.2025.2527536","DOIUrl":"10.1080/07366205.2025.2527536","url":null,"abstract":"Reverse transcription quantitative polymerase chain reaction (RT-qPCR) is widely used for nucleic acid quantification. The use of technical triplicates in RT-qPCR aims to minimize variability and improve reliability but increases reagent consumption, labor, and time. This study systematically evaluates the necessity of technical replicates by analyzing 71,142 cycle threshold (Ct) values from 1,113 RT-qPCR runs across three instruments, two detection chemistries, and 30 operators. Variability within replicates was assessed using metrics such as the coefficient of variation (CV), while the impacts of operator expertise, detection chemistry, instrument calibration, and initial template concentration were explored. The findings challenge the assumption that variability increases at low template concentrations, revealing no correlation between Ct values and CV. While inexperienced operators exhibited slightly higher variability, their replicates were still consistent, with acceptable CVs and low outlier frequencies. Dye-based detection showed greater variability than probe-based. Time since calibration had negligible effects on replicate consistency. Notably, duplicate or single replicates sufficiently approximated triplicate means. These results challenge traditional assumptions about RT-qPCR variability and provide a data-driven framework for optimizing experimental design. This study offers potential for resource savings without compromising data quality, particularly in high-throughput applications or laboratories with limited funds. The data underlying this article are available at https://doi.org/10.5281/zenodo.15072870.","PeriodicalId":8945,"journal":{"name":"BioTechniques","volume":" ","pages":"191-204"},"PeriodicalIF":2.2,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144567032","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

An enzyme-free alcohol-based organoid harvesting solution. 一种无酶酒精类器官收获溶液。

IF 2.2 4区工程技术

BioTechniques Pub Date : 2025-04-01 Epub Date: 2025-07-10 DOI: 10.1080/07366205.2025.2527540

Jimmy Maillard, Lisa Pickard, Udai Banerji

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 : 2025-04-01 Epub 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

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 : 2025-04-01 Epub 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":"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":"153-163"},"PeriodicalIF":2.2,"publicationDate":"2025-04-01","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

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