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Jacqueline Cloos, Jeffrey R Harris, Jeroen J W M Janssen, Angele Kelder, F Huang, Gerrit Sijm, Maike Vonk, Alexander N Snel, Jennifer R Scheick, Willemijn J Scholten, Jannemieke Carbaat-Ham, Dennis Veldhuizen, Diana Hanekamp, Yvonne J M Oussoren-Brockhoff, Gertjan J L Kaspers, Gerrit J Schuurhuis, A Kate Sasser, Gert Ossenkoppele
Response criteria in acute myeloid leukemia (AML) has recently been re-established, with morphologic examination utilized to determine whether patients have achieved complete remission (CR). Approximately half of the adult patients who entered CR will relapse within 12 months due to the outgrowth of residual AML cells in the bone marrow. The quantitation of these remaining leukemia cells, known as minimal or measurable residual disease (MRD), can be a robust biomarker for the prediction of these relapses. Moreover, retrospective analysis of several studies has shown that the presence of MRD in the bone marrow of AML patients correlates with poor survival...
March 5, 2018: Journal of Visualized Experiments: JoVE
Johannes A Eble
The dissociation constant describes the interaction between two partners in the binding equilibrium and is a measure of their affinity. It is a crucial parameter to compare different ligands, e.g., competitive inhibitors, protein isoforms and mutants, for their binding strength to a binding partner. Dissociation constants are determined by plotting concentrations of bound versus free ligand as binding curves. In contrast, titration curves, in which a signal that is proportional to the concentration of bound ligand is plotted against the total concentration of added ligand, are much easier to record...
February 15, 2018: Journal of Visualized Experiments: JoVE
Li Xie, Liqin Chen, Pan Gu, Lanlan Wei, Xuejun Kang
The extraction and analysis of catecholamine neurotransmitters in biological fluids is of great importance in assessing nervous system function and related diseases, but their precise measurement is still a challenge. Many protocols have been described for neurotransmitter measurement by a variety of instruments, including high-pressure liquid chromatography (HPLC). However, there are shortcomings, such as complicated operation or hard-to-detect multiple targets, which cannot be avoided, and presently, the dominant analysis technique is still HPLC coupled with sensitive electrochemical or fluorimetric detection, due to its high sensitivity and good selectivity...
March 1, 2018: Journal of Visualized Experiments: JoVE
Madeline Williams, Smrithi Prem, Xiaofeng Zhou, Paul Matteson, Percy Luk Yeung, Chi-Wei Lu, Zhiping Pang, Linda Brzustowicz, James H Millonig, Emanuel Dicicco-Bloom
Human brain development proceeds through a series of precisely orchestrated processes, with earlier stages distinguished by proliferation, migration, and neurite outgrowth; and later stages characterized by axon/dendrite outgrowth and synapse formation. In neurodevelopmental disorders, often one or more of these processes are disrupted, leading to abnormalities in brain formation and function. With the advent of human induced pluripotent stem cell (hiPSC) technology, researchers now have an abundant supply of human cells that can be differentiated into virtually any cell type, including neurons...
March 2, 2018: Journal of Visualized Experiments: JoVE
Aurélie Fossey-Jouenne, Carine Vergne-Vaxelaire, Anne Zaparucha
Amino alcohols are versatile compounds with a wide range of applications. For instance, they have been used as chiral scaffolds in organic synthesis. Their synthesis by conventional organic chemistry often requires tedious multi-step synthesis processes, with difficult control of the stereochemical outcome. We present a protocol to enzymatically synthetize amino alcohols starting from the readily available L-lysine in 48 h. This protocol combines two chemical reactions that are very difficult to conduct by conventional organic synthesis...
February 16, 2018: Journal of Visualized Experiments: JoVE
Wyatt E Lanik, Lily Xu, Cliff J Luke, Elise Z Hu, Pranjal Agrawal, Victoria S Liu, Rajesh Kumar, Alexa M Bolock, Congrong Ma, Misty Good
Human small intestinal enteroids are derived from the crypts and when grown in a stem cell niche contain all of the epithelial cell types. The ability to establish human enteroid ex vivo culture systems are important to model intestinal pathophysiology and to study the particular cellular responses involved. In recent years, enteroids from mice and humans are being cultured, passaged, and banked away for future use in several laboratories across the world. This enteroid platform can be used to test the effects of various treatments and drugs and what effects are exerted on different cell types in the intestine...
February 15, 2018: Journal of Visualized Experiments: JoVE
Joseph B DeShields, Rachel A Bomberger, James W Woodhall, David L Wheeler, Natalia Moroz, Dennis A Johnson, Kiwamu Tanaka
On-site diagnosis of plant diseases can be a useful tool for growers for timely decisions enabling the earlier implementation of disease management strategies that reduce the impact of the disease. Presently in many diagnostic laboratories, the polymerase chain reaction (PCR), particularly real-time PCR, is considered the most sensitive and accurate method for plant pathogen detection. However, laboratory-based PCRs typically require expensive laboratory equipment and skilled personnel. In this study, soil-borne pathogens of potato are used to demonstrate the potential for on-site molecular detection...
February 23, 2018: Journal of Visualized Experiments: JoVE
Fatima Cabral, Colton M Miller, Katrina M Kudrna, Blake E Hass, Jocelyn G Daubendiek, Brianna M Kellar, Edward N Harris
This protocol demonstrates a method for obtaining high yield and viability for mouse hepatocytes and sinusoidal endothelial cells (SECs) suitable for culturing or for obtaining cell lysates. In this protocol, the portal vein is used as the site for catheterization, rather than the vena cava, as this limits contamination of other possible cell types in the final liver preparation. No special instrumentation is required throughout the procedure. A water bath is used as a source of heat to maintain the temperature of all the buffers and solutions...
February 12, 2018: Journal of Visualized Experiments: JoVE
Miki Dalmau-Pastor, Jordi Vega, Francesc Malagelada, Fernando Peña, Maria Cristina Manzanares-Céspedes
The growing popularity of minimally invasive surgical (MIS) procedures makes it necessary that new anatomical references arise, to aid in tridimensional orientation and localization of structures that are not directly visible to the surgeon. This is especially critical for structures at risk like nerves or blood vessels. Optimization of the handling of cadaveric material and the combination of multiple techniques compensate for the limited availability of adequate specimens. The described protocol combines anatomical plane-by-plane dissection and sectional anatomy of fresh-frozen specimens to help localize relevant structures, such as nerves, arteries, veins and to correctly position the portals during MIS procedures...
February 17, 2018: Journal of Visualized Experiments: JoVE
Barun Das, Jashdeep Bhattacharjee, Preeti, Alaknanda Mishra, Kshama Jain, Srikanth Iyer, Ashwani Kesarwani, Parul Sahu, Prakriti Sinha, Perumal Nagarajan, Pramod Upadhyay
Partial hepatectomy is a versatile and reproducible method to study liver regeneration and the effect of cell based therapeutics in various pathological conditions. Partial hepatectomy also facilitates the increased engraftment and proliferation of transplanted cells by accelerating neovascularization and cell migration towards the liver. Here, we describe a simple protocol for performing 30% hepatectomy and transplantation of cells in the spleen of a non-obese diabetic/severe combined immunodeficient NOD.SCID (NOD...
February 10, 2018: Journal of Visualized Experiments: JoVE
Jonathan T Ting, Brian R Lee, Peter Chong, Gilberto Soler-Llavina, Charles Cobbs, Christof Koch, Hongkui Zeng, Ed Lein
This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have validated the utility of this method for enhancing neuronal preservation and overall brain slice viability. The implementation of this technique by early adopters has facilitated detailed investigations into brain function using diverse experimental applications and spanning a wide range of animal ages, brain regions, and cell types. Steps are outlined for carrying out the protective recovery brain slice technique using an optimized NMDG artificial cerebrospinal fluid (aCSF) media formulation and enhanced procedure to reliably obtain healthy brain slices for patch clamp electrophysiology...
February 26, 2018: Journal of Visualized Experiments: JoVE
Youn Young Shim, Rana Mustafa, Jianheng Shen, Kornsulee Ratanapariyanuch, Martin J T Reaney
Chickpea and other pulses are commonly sold as canned products packed in a thick solution or a brine. This solution has recently been shown to produce stable foams and emulsions, and can act as a thickener. Recently interest in this product has been enhanced through the internet where it is proposed that this solution, now called aquafaba by a growing community, can be used a replacement for egg and milk protein. As aquafaba is both new and being developed by an internet based community little is known of its composition or properties...
February 10, 2018: Journal of Visualized Experiments: JoVE
Andrew K Fuller, Benjamin D Bice, Ashlee R Venancio, Olivia M Crowley, Ambur M Staab, Stephanie J Georges, Julio R Hidalgo, Annika V Warncke, Melinda L Angus-Hill
Several recent studies have illustrated the beneficial effects of living in an enriched environment on improving human disease. In mice, environmental enrichment (EE) reduces tumorigenesis by activating the mouse immune system, or affects tumor bearing animal survival by stimulating the wound repair response, including improved microbiome diversity, in the tumor microenvironment. Provided here is a detailed procedure to assess the effects of environmental enrichment on the biodiversity of the microbiome in a mouse colon tumor model...
February 28, 2018: Journal of Visualized Experiments: JoVE
Grace H Hwang, Jessica L Hopkins, Philip W Jordan
Chromatin spread techniques have been widely used to assess the dynamic localization of various proteins during gametogenesis, particularly for spermatogenesis. These techniques allow for visualization of protein and DNA localization patterns during meiotic events such as homologous chromosome pairing, synapsis and DNA repair. While a few protocols have been described in the literature, general chromatin spread techniques using mammalian prophase oocytes are limited and difficult due to the timing of meiosis initiation in fetal ovaries...
February 26, 2018: Journal of Visualized Experiments: JoVE
Chiara Assunta Pilato, Ilaria Stadiotti, Angela Serena Maione, Valentina Saverio, Valentina Catto, Fabrizio Tundo, Antonio Dello Russo, Claudio Tondo, Giulio Pompilio, Michela Casella, Elena Sommariva
A normal adult heart is composed of several different cell types, among which cardiac mesenchymal stromal cells represent an abundant population. The isolation of these cells offers the possibility of studying their involvement in cardiac diseases, and, in addition, provides a useful primary cell model to investigate biological mechanisms. Here, the method for the isolation of C-MSC from arrhythmogenic cardiomyopathy patients' bioptic samples is described. The endomyocardial biopsy sampling is guided in the right ventricular areas adjacent to the scar visualized by electro-anatomical mapping...
February 28, 2018: Journal of Visualized Experiments: JoVE
Renee Wei-Yan Chow, Paola Lamperti, Emily Steed, Francesco Boselli, Julien Vermot
During embryogenesis, cells undergo dynamic changes in cell behavior, and deciphering the cellular logic behind these changes is a fundamental goal in the field of developmental biology. The discovery and development of photoconvertible proteins have greatly aided our understanding of these dynamic changes by providing a method to optically highlight cells and tissues. However, while photoconversion, time-lapse microscopy, and subsequent image analysis have proven to be very successful in uncovering cellular dynamics in organs such as the brain or the eye, this approach is generally not used in the developing heart due to challenges posed by the rapid movement of the heart during the cardiac cycle...
February 20, 2018: Journal of Visualized Experiments: JoVE
Zhijuan Qiu, Brian S Sheridan
The intestinal immune system plays an essential role in maintaining the barrier function of the gastrointestinal tract by generating tolerant responses to dietary antigens and commensal bacteria while mounting effective immune responses to enteropathogenic microbes. In addition, it has become clear that local intestinal immunity has a profound impact on distant and systemic immunity. Therefore, it is important to study how an intestinal immune response is induced and what the immunologic outcome of the response is...
February 28, 2018: Journal of Visualized Experiments: JoVE
Michael D Guthrie, Donald L Gilbert, David A Huddleston, Ernest V Pedapati, Paul S Horn, Stewart H Mostofsky, Steve W Wu
We describe the development of a reproducible, child-friendly motor response inhibition task suitable for online Transcranial Magnetic Stimulation (TMS) characterization of primary motor cortex (M1) excitability and inhibition. Motor response inhibition prevents unwanted actions and is abnormal in several neuropsychiatric conditions. TMS is a non-invasive technology that can quantify M1 excitability and inhibition using single- and paired-pulse protocols and can be precisely timed to study cortical physiology with high temporal resolution...
February 8, 2018: Journal of Visualized Experiments: JoVE
Ji-Eun Kim, Kyung-Ok Cho
Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status epilepticus (SE) is frequently selected to recapitulate spontaneous recurrent seizures (SRS). Here we present a protocol of SE induction by intraperitoneal (i.p.) injection of pilocarpine and monitoring of chronic recurring seizures in live animals using a wireless telemetry video and electroencephalogram (EEG) system. We demonstrated notable behavioral changes that need attention after pilocarpine injection and their correlation with hippocampal neuronal loss at 7 days and 6 weeks post-pilocarpine...
February 27, 2018: Journal of Visualized Experiments: JoVE
Jing Huang, Peng Guo, Marsha A Moses
The acquisition of the angiogenic phenotype is an essential component of the escape from tumor dormancy. Although several classic in vitro assays (e.g., proliferation, migration, and others) and in vivo models have been developed to investigate and characterize angiogenic and non-angiogenic cell phenotypes, these methods are time and labor intensive, and often require expensive reagents and instruments, as well as significant expertise. In a recent study, we used a novel quantitative phase imaging (QPI) technique to conduct time-lapse and labeling-free characterizations of angiogenic and non-angiogenic human osteosarcoma KHOS cells...
February 16, 2018: Journal of Visualized Experiments: JoVE
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