Articles

A rapid and easy method for the DNA extraction from Cryptococcus neoformans

DNA isolation from C. neoformans is difficult due to a thick and resistant capsule. We have optimized a new and rapid DNA isolation method for Cryptococcus using a short urea treatment followed by a rapid method ...

Authors: Fatma Mseddi, Mohammed Ali Jarboui, Amira Sellami, Hayet Sellami and Ali Ayadi

Quantitative Assessment of Mammary Gland Density in Rodents Using Digital Image Analysis

Rodent models have been used extensively to study mammary gland development and for studies of toxicology and carcinogenesis. Mammary gland gross morphology can visualized via the excision of intact mammary gl...

Authors: John N McGinley and Henry J Thompson

A quantitative PCR method for measuring absolute telomere length

We describe a simple and reproducible method to measure absolute telomere length (aTL) using quantitative real-time polymerase chain reaction (qPCR). This method is based on the Cawthon method for relative mea...

Authors: Nathan J O'Callaghan and Michael Fenech

Biological Procedures Online now publishing with BioMed Central

Authors: Shulin Li

Influence of RT-qPCR Primer Position on EGFR Interference Efficacy in Lung Cancer Cells

Real-time quantitative RT-PCR (RT-qPCR) is a “gold” standard for measuring steady state mRNA levels in RNA interference assays. The knockdown of the epidermal growth factor receptor (EGFR) gene with eight indi...

Authors: Gang Chen, Peter Kronenberger, Erik Teugels and Jacques De Grève

Influence of RT-qPCR primer position on EGFR interference efficacy in lung cancer cells

Real-time quantitative RT-PCR (RT-qPCR) is a "gold" standard for measuring steady state mRNA levels in RNA interference assays. The knockdown of the epidermal growth factor receptor (EGFR) gene with eight indi...

Authors: Gang Chen, Peter Kronenberger, Erik Teugels and Jacques De Grève

Utilization of IκB–EGFP Chimeric Gene as an Indicator to Identify Microbial Metabolites with NF-κB Inhibitor Activity

NF-κB regulates several important expressions, such as cytokine release, anti-apoptosis, adhesion molecule expression, and cell cycle processing. Several NF-κB inhibitors have been discovered as an anti-tumor ...

Authors: Yu-Ling Lin, Yen-Shun Chen, Jui-Hung Hsieh, Ching-Min Lin, Hsin-Yi Wu, Chen-Si Lin, Rea-Min Chu, Kuang-Wen Liao and Yuan-Hsun Hsu

A Method for Serial Tissue Processing and Parallel Analysis of Aberrant Crypt Morphology, Mucin Depletion, and Beta-Catenin Staining in an Experimental Model of Colon Carcinogenesis

The use of architectural and morphological characteristics of cells for establishing prognostic indicators by which individual pathologies are assigned grade and stage is a well-accepted practice. Advances in ...

Authors: John N McGinley, Matthew D Thompson and Henry J Thompson

A Method to Study the Epigenetic Chromatin States of Rare Hematopoietic Stem and Progenitor Cells; MiniChIP–Chip

Dynamic chromatin structure is a fundamental property of gene transcriptional regulation, and has emerged as a critical modulator of physiological processes during cellular differentiation and development. Ana...

Authors: Holger Weishaupt and Joanne L Attema

A New Microsphere-Based Immunoassay for Measuring the Activity of Transcription Factors

There are several traditional and well-developed methods for analyzing the activity of transcription factors, such as EMSA, enzyme-linked immunosorbent assay, and reporter gene activity assays. All of these me...

Authors: Yu-Ling Lin, Yun-Ju Lai, Nu-Man Tsai, Tai-Chu Peng, Yen-Ku Liu, Ru-Ping Lee, Chueh-Jen Tsai and Kuang-Wen Liao

Quick and Simple Detection Technique to Assess the Binding of Antimicrotubule Agents to the Colchicine-Binding Site

Development of antimitotic binding to the colchicine-binding site for the treatment of cancer is rapidly expanding. Numerous antimicrotubule agents are prepared every year, and the determination of their bindi...

Authors: Sébastien Fortin, Jacques Lacroix, Marie-France Côté, Emmanuel Moreau, Éric Petitclerc and René C-Gaudreault

Highly Efficient Stable Expression of Indoleamine 2,3 Dioxygenase Gene in Primary Fibroblasts

Indoleamine 2,3 dioxygenase (IDO) is a potent immunomodulatory enzyme that has recently attracted significant attention for its potential application as an inducer of immunotolerance in transplantation. We hav...

Authors: Alireza Moeen Rezakhanlou, Darya Habibi, Amy Lai, Reza B Jalili, Christopher J Ong and Aziz Ghahary

Optimized RNA Extraction and Northern Hybridization in Streptomycetes

Northern blot hybridization is a useful tool for analyzing transcript patterns. To get a picture of what really occurs in vivo, it is necessary to use a protocol allowing full protection of the RNA integrity a...

Authors: Marcello Tagliavia, Anna Taravella, Sandra Marineo, Anna Maria Puglia and Mario La Farina

Collection of Epithelial Cells from Rodent Mammary Gland Via Laser Capture Microdissection Yielding High-Quality RNA Suitable for Microarray Analysis

Laser capture microdissection (LCM) enables collection of cell populations highly enriched for specific cell types that have the potential of yielding critical information about physiological and pathophysiolo...

Authors: John N McGinley, Zongjian Zhu, Weiqin Jiang and Henry J Thompson

Comparison between Culture Conditions Improving Growth and Differentiation of Blood and Bone Marrow Cells Committed to the Endothelial Cell Lineage

The aim of this study was to compare different cell sources and culture conditions to obtain endothelial progenitor cells (EPCs) with predictable antigen pattern, proliferation potential and in vitro vasculoge...

Authors: Claudio Muscari, Chiara Gamberini, Ilaria Basile, Francesca Bonafé, Simond Valgimigli, Ombretta Capitani, Carlo Guarnieri and Claudio Marcello Caldarera

Ocular Proteomics with Emphasis on Two-Dimensional Gel Electrophoresis and Mass Spectrometry

The intention of this review is to provide an overview of current methodologies employed in the rapidly developing field of ocular proteomics with emphasis on sample preparation, two-dimensional polyacrylamide...

Authors: Nakul Mandal, Steffen Heegaard, Jan Ulrik Prause, Bent Honoré and Henrik Vorum

A Novel mRNA Level Subtraction Method for Quick Identification of Target-Orientated Uniquely Expressed Genes Between Peanut Immature Pod and Leaf

Subtraction technique has been broadly applied for target gene discovery. However, most current protocols apply relative differential subtraction and result in great amount clone mixtures of unique and differe...

Authors: Ning Wu, Kanyand Matand and Sonya Williams

Change in Hemoglobin Levels due to Anesthesia in Mice: An Important Confounder in Studies on Hematopoietic Drugs

Analgesic and anesthetic drugs may have an impact on the results achieved from animal experiments. In the study presented here, we try to enlighten whether anesthesia with fentanyl/fluniasone and midazolam (Hy...

Authors: Anita Gothelf, Pernille Hojman and Julie Gehl

Critical Appraisal of the MTT Assay in the Presence of Rottlerin and Uncouplers

Rottlerin is a natural product isolated from Mallotus philippinensis. This polyphenolic compound, originally described as a selective inhibitor of PKCδ, can inhibit many other PKC-unrelated kinases and has a numb...

Authors: Emanuela Maioli, Claudia Torricelli, Vittoria Fortino, Filippo Carlucci, Valentina Tommassini and Adriana Pacini

A Modified Coupled Enzyme Method for O-linked GlcNAc Transferase Activity Assay

In order to determine the activity of O-linked GlcNAc transferase (OGT), a modified coupled enzyme method was proposed. This method was based on the measurement of uridine 5'-(trihydrogen diphosphate) (UDP), a...

Authors: Lianwen Zhang, Feifei Ren, Jing Li, Xiaofeng Ma and Peng Wang

A Practical Guide to Rodent Islet Isolation and Assessment

Pancreatic islets of Langerhans secrete hormones that are vital to the regulation of blood glucose and are, therefore, a key focus of diabetes research. Purifying viable and functional islets from the pancreas...

Authors: Jeffrey D Carter, Stacey B Dula, Kathryn L Corbin, Runpei Wu and Craig S Nunemaker

Dynamic Monitoring of Cellular Remodeling Induced by the Transforming Growth Factor-β1

The plasticity of differentiated adult cells could have a great therapeutic potential, but at the same time, it is characteristic of progression of serious pathological states such as cancer and fibrosis. In t...

Authors: Andrea Staršíchová, Lukáš Kubala, Eva Lincová, Zuzana Pernicová, Alois Kozubík and Karel Souček

The Possible Cellular Mechanism for Extending Lifespan of Mice with Rapamycin

Authors: Shulin Li

MicroRNA Expression Profiling by Bead Array Technology in Human Tumor Cell Lines Treated with Interferon-Alpha-2a

MicroRNAs are positive and negative regulators of eukaryotic gene expression that modulate transcript abundance by specific binding to sequence motifs located prevalently in the 3' untranslated regions of targ...

Authors: Fredy Siegrist, Thomas Singer and Ulrich Certa

Segmentation of Extrapulmonary Tuberculosis Infection Using Modified Automatic Seeded Region Growing

In the image segmentation process of positron emission tomography combined with computed tomography (PET/CT) imaging, previous works used information in CT only for segmenting the image without utilizing the i...

Authors: Iman Avazpour, M Iqbal Saripan, Abdul Jalil Nordin and Rajaa Syamsul Azmir Iman Abdullah

Lentiviral-Mediated Transgene Expression Can Potentiate Intestinal Mesenchymal-Epithelial Signaling

Mesenchymal-epithelial signaling is essential for the development of many organs and is often disrupted in disease. In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effecti...

Authors: Adria D Dismuke, Aimee D Kohn, Randall T Moon and Melissa H Wong

A Modified Protocol for Bisulfite Genomic Sequencing of Difficult Samples

The bisulfite genomic sequencing protocol is a widely used method for analyzing DNA methylation. It relies on the deamination of unmethylated cytosine residues to uracil; however, its high rates of DNA degrada...

Authors: Jane J Pappas, André Toulouse and WEC Bradley

A Method for Structure–Activity Analysis of Quorum-Sensing Signaling Peptides from Naturally Transformable Streptococci

Many species of streptococci secrete and use a competence-stimulating peptide (CSP) to initiate quorum sensing for induction of genetic competence, bacteriocin production, and other activities. These signaling...

Authors: XiaoLin Tian, Raymond T Syvitski, TianLei Liu, Nadine Livingstone, David L Jakeman and Yung-Hua Li

A Framework for White Blood Cell Segmentation in Microscopic Blood Images Using Digital Image Processing

Evaluation of blood smear is a commonly clinical test these days. Most of the time, the hematologists are interested on white blood cells (WBCs) only. Digital image processing techniques can help them in their...

Authors: Farnoosh Sadeghian, Zainina Seman, Abdul Rahman Ramli, Badrul Hisham Abdul Kahar and M-Iqbal Saripan

SDS-PAGE-Based Quantitative Assay for Screening of Kidney Stone Disease

Kidney stone disease is a common health problem in industrialised nations. We developed a SDS-PAGE-based method to quantify Tamm Horsfall glycoprotein (THP) for screening of kidney stone disease. Urinary prote...

Authors: Lau Wai-Hoe, Leong Wing-Seng, Zhari Ismail and Gam Lay-Harn

Size and Shape of Protein Molecules at the Nanometer Level Determined by Sedimentation, Gel Filtration, and Electron Microscopy

An important part of characterizing any protein molecule is to determine its size and shape. Sedimentation and gel filtration are hydrodynamic techniques that can be used for this medium resolution structural ...

Authors: Harold P Erickson

In Vivo Imaging of Far-red Fluorescent Proteins after DNA Electrotransfer to Muscle Tissue

DNA electrotransfer to muscle tissue yields long-term, high levels of gene expression; showing great promise for future gene therapy. We want to characterize the novel far-red fluorescent protein Katushka as a...

Authors: Pernille Hojman, Jens Eriksen and Julie Gehl

Detection and Processing Techniques of FECG Signal for Fetal Monitoring

Fetal electrocardiogram (FECG) signal contains potentially precise information that could assist clinicians in making more appropriate and timely decisions during labor. The ultimate reason for the interest in...

Authors: MA Hasan, MBI Reaz, MI Ibrahimy, MS Hussain and J Uddin

Computational Biology Methods and Their Application to the Comparative Genomics of Endocellular Symbiotic Bacteria of Insects

Comparative genomics has become a real tantalizing challenge in the postgenomic era. This fact has been mostly magnified by the plethora of new genomes becoming available in a daily bases. The overwhelming lis...

Authors: Jennifer Commins, Christina Toft and Mario A Fares

Comparison of Enzymatic and Non-Enzymatic Means of Dissociating Adherent Monolayers of Mesenchymal Stem Cells

The dissociation of adherent mesenchymal stem cell (MSC) monolayers with trypsin and enzyme-free dissociation buffer was compared. A significantly lower proportion of viable cells were obtained with enzyme-fre...

Authors: Boon C Heng, Catherine M Cowan and Shubhayu Basu

Competitive RT-PCR Strategy for Quantitative Evaluation of the Expression of Tilapia (Oreochromis niloticus) Growth Hormone Receptor Type I

Quantization of gene expression requires that an accurate measurement of a specific transcript is made. In this paper, a quantitative reverse transcription-polymerase chain reaction (RT-PCR) by competition for...

Authors: Alina Rodríguez-Mallon, Yamilet Cárdenas, Juana María Lugo, Aymé Oliva, Antonio Morales and Mario Pablo Estrada

Streptozotocin, Type I Diabetes Severity and Bone

As many as 50% of adults with type I (T1) diabetes exhibit bone loss and are at increased risk for fractures. Therapeutic development to prevent bone loss and/or restore lost bone in T1 diabetic patients requi...

Authors: Katherine Motyl and Laura R McCabe

Ultra-Fast and Optimized Method for the Preparation of Rodent Testicular Cells for Flow Cytometric Analysis

Homogeneity of cell populations is a prerequisite for the analysis of biochemical and molecular events during male gamete differentiation. Given the complex organization of the mammalian testicular tissue, var...

Authors: Rosana Rodríguez-Casuriaga, Adriana Geisinger, Beatriz López-Carro, Valentina Porro, Rodolfo Wettstein and Gustavo A Folle

A method to generate human mesenchymal stem cell-derived neurons which express and are excited by multiple neurotransmitters

The present study describes a protocol to generate heterogenous populations of neurotransmitter-producing neurons from human mesenchymal stem cells (MSCs). MSCs are bone marrow (BM)-derived cells which undergo...

Authors: Steven J. Greco, Chunyi Zhou, Jiang-Hong Ye and Pranela Rameshwar

A rapid microwell fluorescence immunoassay for cellular protein detection

In this paper, we describe a simple, rapid, specific, sensitive, and reliable method, the FICP method (Fluorescence Immunoassay for Cellular Protein detection) which is readily applicable to the detection of p...

Authors: Carole Lavigne, Armèle de Guignée and Alain R. Thierry

Tracking and ablating subpopulations of epiblast cells in the chick embryo

The early chick embryo contains subpopulations of cells that express lineage-specific transcription factors. We have developed protocols to examine the role of these cells during development that involve label...

Authors: Jacquelyn Gerhart, Christine Neely, Jessica Pfautz and Mindy George-Weinstein

Characterizing gene family evolution

Gene families are widely used in comparative genomics, molecular evolution, and in systematics. However, they are constructed in different manners, their data analyzed and interpreted differently, with differe...

Authors: David A. Liberles and Katharina Dittmar

Cell culture-based analysis of postsynaptic membrane assembly in muscle cells

We report a method for studying postsynaptic membrane assembly utilizing the replating of aneural cultures of differentiated skeletal muscle cells onto laminin-coated surfaces. A significant limitation to the ...

Authors: Getu Teressa and Joav Prives

A method for purification, identification and validation of DNMT1 mRNA binding proteins

DNA methyltransferase 1 (DNMT1) is the enzyme responsible for the maintenance of DNA methylation patterns during cell division. DNMT1 expression is tightly regulated within the cell cycle. Our previous study s...

Authors: Alexander Unterberger, Jérôme Torrisani and Moshe Szyf

Optimized two-dimensional thin layer chromatography to monitor the intracellular concentration of acetyl phosphate and other small phosphorylated molecules

Acetyl phosphate (acetyl-P) serves critical roles in coenzyme A recycling and ATP synthesis. It is the intermediate of the Pta-AckA pathway that inter-converts acetyl-coenzyme A and acetate. Acetyl-P also can ...

Authors: David H. Keating, Ana Shulla, Adam H. Klein and Alan J. Wolfe

Identification of a calcitriol-regulated Sp-1 site in the promoter of human CD14 using a combined western blotting elect rophoresis mobility shift assay (WEMSA)

Calcitriol (1α, 25-dihydroxyvitamin D₃) induces the expression of CD14 in mononuclear phagocytes. The mechanisms accounting for this have been unclear since the promoter of CD14 does not contain a canonical vitam...

Authors: Alireza Moeenrezakhanlou, Devki Nandan and Neil E. Reiner

Quantitative evaluation of signaling events in Drosophila S2 cells

Drosophila activates a robust defense response to gram-negative bacteria through the Immune deficiency (Imd) pathway. Imd signaling proceeds through c-Jun N-terminal Kinase (JNK), NF-kB and caspa...

Authors: David Bond, David A. Primrose and Edan Foley

Recent advances in freeze-fracture electron microscop: the replica immunolabeling technique

Freeze-fracture electron microscopy is a technique for examining the ultrastructure of rapidly frozen biological samples by transmission electron microscopy. Of a range of approaches to freeze-fracture cytoche...

Authors: Horst Robenek and Nicholas J. Severs

In vivo and in vitro techniques for comparative study of antiviral T-cell responses in the amphibian Xenopus

Activation of lymphocytes in mammals is often quantified by measuring the amount of proliferation during the expansion phase of an immune response. Bromodeoxyuridine (BrdU) incorporation and carboxyfluorescein...

Authors: Heidi Morales and Jacques Robert

Rapid sitrapid site-directed domain scanning mutagenesis of enteropathogenic Escherichia coli espD

We developed a rapid mutagenesis method based on a modification of the QuikChange® system (Stratagene) to systemically replace endogenous gene sequences with a unique similar size sequence tag. The modificatio...

Authors: Qiwen Deng, Wensheng Luo and Michael S. Donnenberg