Protocols

• Books for Xenopus Research and Protocols
  • Normal Table of Xenopus Laevis (Daudin), Nieuwkoop and Faber (Third edition), 1994 [1]
    • a systematic and chronological description of Xenopus laevis development
  • Early Development of Xenopus laevis: A Laboratory Manual, Sive, Grainger, and Harland, 2000 [2]
    • comprehensive collection of protocols for the study of early development in Xenopus embryos
  • The Laboratory Xenopus sp., Green, 2010 [3]
    • a highly detailed manual containing Xenopus husbandry, management, veterinary care, and frog and equipment vendor information
  • Xenopus Protocols: Cell Biology and Signal Transduction, Liu (First edition), 2006 [4]
    • step-by-step laboratory instructions, an introduction outlining the principles behind the technique, lists of the necessary equipment and reagents
  • Xenopus laevis: Practical Uses in Cell and Molecular Biology, Volume 36, Kay and Peng, 1991 [5]
    • detailed description of a wide range of uses for Xenopus laevis oocytes and embryos in cell and molecular biology
  • Transgenic Xenopus: Microinjection Methods and Developmental Neurobiology, Seidman and Soreq, 1997 [6]
    • referenced guide to the use of microinjected embryos studying the role and regulation of nervous system proteins during development
  • Atlas of Xenopus Development, Bernardini, Prati, Bonetti, and Scari, 1999 [7]
    • scanning, transmission, and light microscopy images of Xenopus embryonic development
  • The early development of Xenopus laevis. An atlas of the histology, Hausen and Riebesell, 1991 [8]
    • detailed histological sections of Xenopus embryonic development
  • Color Atlas of Xenopus laevis Histology, Wiechmann and Wirsig-Wiechmann, 2003 [9]
    • central source on the microscopic anatomy of cells, tissues, and major organs of Xenopus laevis

• Online Resources for Xenopus Research and Protocols
  • Cold Spring Harbor Protocols - Xenopus [10], subscription required
  • The Journal of Visualized Experiments (JOVE) - Xenopus embryo [11] - excellent video demonstrations
    • Production of Transgenic Xenopus laevis by Restriction Enzyme Mediated Integration and Nuclear Transplantation - Enrique Amaya & Kristen Kroll [12]
    • Tissue Determination Using the Animal Cap Transplant (ACT) Assay in Xenopus laevis - Andrea S. Viczian & Michael E. Zuber [13]
    • Blastomere Explants to Test for Cell Fate Commitment During Embryonic Development - Paaqua A. Grant, Mona B. Herold & Sally A. Moody [14]
    • Neural Explant Cultures from Xenopus laevis - Laura Anne Lowery, Anna E.R. Faris, Alina Stout & David Van Vactor [15]
    • Polarized Translocation of Fluorescent Proteins in Xenopus Ectoderm in Response to Wnt Signaling - Keiji Itoh & Sergei Y. Sokol [16]
    • Production of Xenopus tropicalis Egg Extracts to Identify Microtubule-associated RNAs - Judith A. Sharp & Mike D. Blower [17]
    • Electroporation of Craniofacial Mesenchyme - Jacqueline M. Tabler & Karen J. Liu [18]
    • Simultaneous Pre- and Post-synaptic Electrophysiological Recording from Xenopus Nerve-muscle Co-cultures - Bruce Yazejian, Rita M. Yazejian, Rachel Einarsson & Alan D. Grinnell [19]
    • Dissection, Culture, and Analysis of Xenopus laevis Embryonic Retinal Tissue - Molly J. McDonough, Chelsea E. Allen, Ng-Kwet-Leok A. Ng-Sui-Hing, Brian A. Rabe, Brittany B. Lewis & Margaret S. Saha [20]
    • Live-cell Imaging and Quantitative Analysis of Embryonic Epithelial Cells in Xenopus laevis - Sagar D. Joshi & Lance A. Davidson [21]
    • Plastic Embedding and Sectioning of Xenopus laevis Embryos - Souichi Ogata1, Shimako Kawauchi, Anne Calof & Ken W.Y. Cho [22]
    • Dissection of Organizer and Animal Pole Explants from Xenopus laevis Embryos and Assembly of a Cell Adhesion Assay - Souichi Ogata & Ken W.Y. Cho [23]
    • Fertilization of Xenopus oocytes using the Host Transfer Method - Patricia N. Schneider, Alissa M. Hulstrand & Douglas W. Houston [24]
    • Microinjection of Xenopus Laevis Oocytes - Sarah Cohen, Shelly Au & Nelly Panté - [25]
    • Comparative in vivo Study of gp96 Adjuvanticity in the Frog Xenopus laevis - Hristina Nedelkovska, Tanya Cruz-Luna, Pamela McPherson & Jacques Robert [26]
    • Two Types of Assays for Detecting Frog Sperm Chemoattraction - Lindsey A. Burnett, Nathan Tholl & Douglas E. Chandler [27]
    • Study of the DNA Damage Checkpoint using Xenopus Egg Extracts - Jeremy Willis, Darla DeStephanis, Yogin Patel, Vrushab Gowda & Shan Yan [28]
    • Patch Clamp Recording of Ion Channels Expressed in Xenopus Oocytes - Austin L Brown, Brandon E. Johnson & Miriam B. Goodman [29]
    • Measuring Cation Transport by Na,K- and H,K-ATPase in Xenopus Oocytes by Atomic Absorption Spectrophotometry: An Alternative to Radioisotope Assays - Katharina L. Dürr, Neslihan N. Tavraz, Susan Spiller & Thomas Friedrich [30]
    • Preparation and Fractionation of Xenopus laevis Egg Extracts - Marie K. Cross & Maureen Powers [31]
    • Obtaining Eggs from Xenopus laevis Females - Marie K. Cross & Maureen Powers [32]
    • In Vitro Nuclear Assembly Using Fractionated Xenopus Egg Extracts - Marie Cross & Maureen Powers [33]
    • Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in Xenopus oocytes - Junqiu Yang, Kelli Delaloye, Urvi S. Lee & Jianmin Cui [34]
    • Visualizing RNA Localization in Xenopus Oocytes - James A. Gagnon & Kimberly L. Mowry [35]
    • Single Cell Electroporation in vivo within the Intact Developing Brain - D. Sesath Hewapathirane & Kurt Haas - [36]

• Animal Husbandry
  • obtain frogs and frog housing equipment [37]
  • Xenopus Express links [38] - Xenopus Express husbandry [39]
  • Xenopus tropicalis and Xenopus laevis environmental parameter standard operating procedure (Kroll lab)

• Lab Solutions and Reagents - click to view expanded list
  • culture media
  • immunohistochemistry
  • in situ hybridization
  • fixatives
  • DNA/RNA preparation
  • embryo manipulation/dissection
  • microinjection
  • general buffers and salt solutions

• Generating Embryos
  • inducing egg laying via hCG injection (Vize lab)
  • anesthesia, in vitro fertilization, and natural mating of Xenopus tropicalis (Conlon lab)
  • benzocaine stock, 100x (Khokha lab)
  • in vitro fertilization of Xenopus laevis (Conlon lab)
  • high-salt solution for egg laying (Zorn lab/Ron Laskey)
  • dejellying embryos (Sive et al. 2000)
  • dejellying embryos (Zorn lab)

• Transgenics
  • transgenesis in Xenopus laevis - adapted from Amaya et al 2001 (Conlon lab)
  • tadpole tail sampling for genomic DNA (Conlon lab)
  • Production of transgenic Xenopus laevis by restriction enzyme mediated integration and nuclear transplantation. Amaya E, Kroll K. J Vis Exp. 2010 Aug 21;(42). [40]
  • A method for generating transgenic frog embryos. Ishibashi S, Kroll KL, Amaya E. Methods Mol Biol. 2008;461:447-66. [41]
  • A method for generating transgenic frog embryos. Amaya E, Kroll KL. Methods Mol Biol. 1999;97:393-414. [42]

• In Situ Hybridization
  • in situ hybridization, brief version (Harland lab)
  • whole-mount in situ hybridization (Conlon lab)
    • in situ checklist (Conlon lab)
  • fluorescent in-situs and FCIS (Vize lab)
    • Wholemount embryo FISH / Double FISH
    • Flourescin Tyramide Synthesis
    • Cy3 Tyramide Synthesis
    • FCIS

• Immunohisochemistry
  • wholemount antibody staining (CSH Protocols) (Wallingford lab)
  • wholemount antibody staining - alkaline phosphatase detection (Conlon lab)
  • wholemount antibody staining - adapted from Dan Weeks' protocol (Conlon lab)
  • immunostaining sectioned embryos (Conlon lab)
  • hydrochloric acid antigen retrieval (Zorn lab)

• Histology
  • cryosectioning Xenopus embryos (Conlon lab)

• Other Embryo Staining Protocols
  • benzidine-peroxidase staining for blood (Conlon lab)
  • benzidine-peroxidase staining for blood (Thomsen lab)
  • apoptosis via TUNEL (Veenstra lab)
  • TUNEL staining of whole Xenopus embryos - Harland protocol (Conlon lab)

• Other Immuno Protocols and Protein Protocols
  • immunoprecipitation (Stukenberg lab)
  • immunofluorescence with biotinylated antibodies (Stukenberg lab)
  • Purifying Polyclonal Antibodies using Affinity Columns/Chromatography
    • Making an Affinity Column
    • Running an Affinity Column
  • whole embryo protein extraction with freon (Conlon lab)
  • SDS-PAGE gels (Conlon lab)
  • western blot protocol (Conlon lab)
  • silver staining protein gels (Stukenberg lab)

• Nucleic Acid Protocols
  • RNA Isolation
    • Preparation of RNA Using Proteinase K/LiCl (Sive et al. 2000)
    • Preparation of RNA (In Vitro Transcription) (Sive et al. 2000)
  • DNA Isolation
    • Extraction of DNA from Single Embryos (Sive et al. 2000)
    • Isolation of DNA from Red Blood Cells (Sive et al. 2000)

• Oocyte Transfer Technique (Heasman/Wylie labs)
  • Part 1: Choosing Oligodeoxyribonucleotides (ODNs)
  • Part 2: ODN Modification
  • Part 3: Host Transfer Technique
  • Part 4: Fertilization and Development

• Xenopus Oocyte and Egg Extracts
  • crude interphase egg extract (Shechter lab)
  • isolation of chromatin from egg extract and histone recovery (Shechter lab)
  • preparation of cycling egg extract (Shechter lab)
  • cycling extracts, mike's protocol (Stukenberg lab)
  • cycling extracts, sara's protocol (Stukenberg lab)
  • CSF extract, mike's protocol (Stukenberg lab)
  • CSF extract, sara's protocol (Stukenberg lab)
  • assaying H1 kinase in extracts (Stukenberg lab)

• Xenopus Tissue Culture
  • tissue culture of Xenopus cell lines S3, XTC (Stukenberg lab)
  • isolation of mitotic chromosomes from XTC cells (Stukenberg lab)
  • in-vitro kinetochore-microtubule binding assays (Stukenberg lab)