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Top 10 Most-read Protocols

Most Read rankings are calculated based on published articles (since 2015) read in the last two months. Rankings are calculated monthly. Rankings are based on page view data provided by Google Analytics.

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Quantification of Mouse Hematopoietic Progenitors’ Formation Using Time-lapse Microscopy and Image Analysis

Multiple Stepwise Gene Knockout Using CRISPR/Cas9 in Escherichia coli

Authors: Enrico König, Francesca Zerbini, Ilaria Zanella, Davide Fraccascia and Guido Grandi, date: 01/20/2018, view: 6673, Q&A: 1

With the recent implementation of the CRISPR/Cas9 technology as a standard tool for genome editing, laboratories all over the world are undergoing one of the biggest advancements in molecular biology since PCR. The key advantage of this method is its simplicity and universal applicability for species of any phylum. Of particular interest is the ...

Measurement of Oxygen Consumption Rate (OCR) and Extracellular Acidification Rate (ECAR) in Culture Cells for Assessment of the Energy Metabolism

Authors: Birte Plitzko and Sandra Loesgen, date: 05/20/2018, view: 8701, Q&A: 0

Mammalian cells generate ATP by mitochondrial (oxidative phosphorylation) and non-mitochondrial (glycolysis) metabolism. Cancer cells are known to reprogram their metabolism using different strategies to meet energetic and anabolic needs (Koppenol et al., 2011; Zheng, 2012). Additionally, each cancer tissue has its own individual ...

Sulforhodamine B (SRB) Assay in Cell Culture to Investigate Cell Proliferation

Authors: Esteban A. Orellana and Andrea L. Kasinski, date: 11/05/2016, view: 22202, Q&A: 0

The SRB assay has been used since its development in 1990 (Skehan et al., 1990) to inexpensively conduct various screening assays to investigate cytotoxicity in cell based studies (Vichai and Kirtikara, 2006). This method relies on the property of SRB, which binds stoichiometrically to proteins under mild acidic conditions and then can be ...

BODIPY 493/503 Staining of Neutral Lipid Droplets for Microscopy and Quantification by Flow Cytometry

Authors: Bo Qiu and M. Celeste Simon, date: 09/05/2016, view: 20272, Q&A: 0

Lipid droplets (LDs) are ubiquitous, dynamic organelles and function as a storage depot for neutral lipids, including triglycerides and cholesterol esters (Walther and Farese, 2012). The movement of lipid species into and out of LDs impacts a variety of cellular processes, such as energy homeostasis, lipid-based signaling, and membrane homeostasis ...

Analysis of Protein Stability by the Cycloheximide Chase Assay

Authors: Shih-Han Kao, Wen-Lung Wang, Chi-Yuan Chen, Yih-Leong Chang, Yi-Ying Wu, Yi-Ting Wang, Shu-Ping Wang, Alexey I Nesvizhskii, Yu-Ju Chen, Tse-Ming Hong and Pan-Chyr Yang, date: 01/05/2015, view: 39403, Q&A: 1

Comparison of protein stability in eukaryotic cells has been achieved by cycloheximide, which is an inhibitor of protein biosynthesis due to its prevention in translational elongation. It is broadly used in cell biology in terms of determining the half-life of a given protein and has gained much popularity in cancer research. Here we present a ...

Cell Synchronization by Double Thymidine Block

Authors: Guo Chen and Xingming Deng, date: 09/05/2018, view: 3577, Q&A: 1

Cell synchronization is widely used in studying mechanisms involves in regulation of cell cycle progression. Through synchronization, cells at distinct cell cycle stage could be obtained. Thymidine is a DNA synthesis inhibitor that can arrest cell at G1/S boundary, prior to DNA replication. Here, we present the protocol to synchronize cells at ...

Superoxide Dismutase (SOD) and Catalase (CAT) Activity Assay Protocols for Caenorhabditis elegans

Authors: Jing Zhang, Rui Chen, Zhenyang Yu and Lili Xue, date: 08/20/2017, view: 9482, Q&A: 0

Assays for superoxide dismutase (SOD) and catalase (CAT) activities are widely employed to indicate antioxidant responses underlying the toxic effects of test chemicals. Yet, earlier studies mainly described the procedures as performed according to manufacturer’s instructions without modifications that are specific to any organisms. The present ...

Dot Blot Analysis of N⁶-methyladenosine RNA Modification Levels

Authors: Lisha Shen, Zhe Liang and Hao Yu, date: 01/05/2017, view: 7658, Q&A: 2

N⁶-methyladenosine (m⁶A) is the most prevalent internal modification of eukaryotic messenger RNA (mRNA). The total amount of m⁶A can be detected by several methods, such as dot blot analysis using specific m⁶A antibodies and quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) (Fu et al ...

Hemagglutination Inhibition (HI) Assay of Influenza Viruses with Monoclonal Antibodies

Authors: Ying Wu, MyungSam Cho, David Shore, Manki Song, JungAh Choi, Tao Jiang, Yong-Qiang Deng, Melissa Bourgeois, Lynn Almli, Hua Yang, Li-Mei Chen, Yi Shi, Jianxu Qi, An Li, Kye Sook Yi, MinSeok Chang, Jin Soo Bae, HyunJoo Lee, JiYoung Shin, James Stevens, SeoungSuh Hong, Cheng-Feng Qin, George F. Gao, Shin Jae Chang and Ruben O. Donis, date: 06/05/2016, view: 13740, Q&A: 0

Heamagglutination is inhibited when antibodies are present because antibodies to the influenza virus will prevent attachment of the virus to red blood cells. The highest dilution of antibody that prevents hemagglutination is called the HI titer. Human monoclonal antibodies generated from single human B cells were tested to characterize their ...

Isolation of PBMCs Using Vacutainer^® Cellular Preparation Tubes (CPT^TM)

Authors: Alaina Puleo, Chantia Carroll, Holden Maecker and Rohit Gupta, date: 01/20/2017, view: 12776, Q&A: 0

Peripheral blood mononuclear cell (PBMC) isolation is commonly done via density gradient centrifugation over Ficoll-Hypaque, a labor-intensive procedure that requires skilled technicians and can contribute to sample variability. Cellular Preparation Tubes (CPTs) are Vacutainer blood draw tubes that contain Ficoll-Hypaque and a gel plug that ...

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