Protocols

Books for Xenopus Research and Protocols

• Xenopus Protocols: Post-Genomic Approaches. Hoppler and Vize, 2012 [1]
  • expanded second edition with novel approaches inspired by X. tropicalis genome sequencing.

• The Laboratory Xenopus sp. Green, 2010 [2]
  • 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 [3]
  • step-by-step laboratory instructions, an introduction outlining the principles behind the technique, lists of the necessary equipment and reagents.

• Color Atlas of Xenopus laevis Histology, Wiechmann and Wirsig-Wiechmann, 2003 [4]
  • central source on the microscopic anatomy of cells, tissues, and major organs of Xenopus laevis.

• Early Development of Xenopus laevis: A Laboratory Manual, Sive, Grainger, and Harland, 2000 [5]
  • comprehensive collection of protocols for the study of early development in Xenopus embryos

• 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.

• Normal Table of Xenopus laevis (Daudin), Nieuwkoop and Faber (Third edition), 1994 [8]
  • a systematic and chronological description of Xenopus laevis development.

• Xenopus laevis: Practical Uses in Cell and Molecular Biology, Volume 36, Kay and Peng, 1991 [9]
  • detailed description of a wide range of uses for Xenopus laevis oocytes and embryos in cell and molecular biology.

• The early development of Xenopus laevis. An atlas of the histology, Hausen and Riebesell, 1991 [10]
  • detailed histological sections of Xenopus embryonic development

Xenopus Protocols and Video Demostrations: Online Resources

Journal of Visualized Experiments (JOVE) - Xenopus embryo [11] - excellent video demonstrations and detailed protocols for a wide variety of lab protocols using Xenopus

• • Functional Evaluation of Olfactory Pathways in Living Xenopus Tadpoles - Terni et al. -[12]
  • Obtaining Eggs from Xenopus laevis Females - Cross & Powers [13]
  • Microinjection of Xenopus laevis Oocytes - Cohen et al. - [14]
  • Fertilization of Xenopus oocytes, Host Transfer Method - Schneider et al. [15]
  • Tissue Determination Using Animal Cap Transplant (ACT) Assay in X. laevis - Viczian & Zuber [16]
  • Blastomere Explants to Test for Cell Fate Commitment - Grant et al. [17]
  • Transgenic Xenopus laevis by Restriction Enzyme Mediated Integration and Nuclear Transplantation - Amaya & Kroll [18]
  • Organizer and Animal Pole Explants from X. laevis Embryos and Cell Adhesion Assay - Ogata & Cho [19]
  • Plastic Embedding and Sectioning of X. laevis Embryos - Ogata1 et al. [20]
  • Visualizing RNA Localization in Xenopus Oocytes - Gagnon & Mowry [21]
  • Neural Explant Cultures from Xenopus laevis - Lowery et al. [22]
  • Translocation of Fluorescent Proteins in Xenopus Ectoderm in Response to Wnt Signaling - Itoh & Sokol [23]
  • Dissection, Culture, and Analysis of X. laevis Embryonic Retinal Tissue - McDonough et al. [24]
  • X. tropicalis Egg Extracts to Identify Microtubule-associated RNAs - Sharp & Blower [25]
  • Electroporation of Craniofacial Mesenchyme - Tabler & Liu [26]
  • Electrophysiological Recording from Xenopus Nerve-muscle Co-cultures - Yazejian et al. [27]
  • Single Cell Electroporation in vivo within the Intact Developing Brain - Hewapathirane & Haas - [28]
  • Live-cell Imaging and Quantitative Analysis of Embryonic Epithelial Cells - Joshi & Davidson [29]
  • Preparation and Fractionation of Xenopus laevis Egg Extracts - Cross & Powers [30]
  • In Vitro Nuclear Assembly Using Fractionated Xenopus Egg Extracts - Cross & Powers [31]
  • Study of the DNA Damage Checkpoint using Xenopus Egg Extracts - Willis et al. [32]
  • Two Types of Assays for Detecting Frog Sperm Chemoattraction - Burnett et al. [33]
  • Comparative in vivo Study of gp96 Adjuvanticity in X. laevis - Nedelkovska et al. 2010. [34]
  • Patch Clamp and Perfusion Techniques - Yang et al. [35]
  • Patch Clamp Recording of Ion Channels - Brown et al. [36]
  • Cation Transport in Xenopus Oocytes - Dürr et al.[37]
  • Stem cell-like Xenopus Embryonic Explants to Study Early Neural Developmental Features In Vitro and In Vivo - BC.Durand [38]
  • Dissection of Xenopus laevis Neural Crest for in vitro Explant Culture or in vivo Transplantation - Millet & Monsoro-Burq [39]
  • Microinjection of DNA into Eyebuds in Xenopus laevis Embryos and Imaging of GFP Expressing Optic Axonal Arbors in Intact, Living Xenopus Tadpoles- Dao, Jones and Elul. 2019. [40]
  • Reconstitution Of β-catenin Degradation In Xenopus Egg Extract. Chen et al. 2020. [41]

• • Xenopus laevis Egg Extract Preparation and Live Imaging Methods for Visualizing Dynamic Cytoplasmic Organization. Xianrui Cheng and James Farrell. 2021. [42]

Protocols published in non-CSHL Journals and from Xenopus labs -click to view-

Xenopus Sperm cryopreservation

• • Transitioning from a research protocol to a scalable applied pathway for Xenopus laevis sperm cryopreservation at a national stock center: The effect of cryoprotectants. Lucía Arregui, Jack C. Koch, Terrence R. Tiersch. 2023. JEZ-B molecular and Developmental Evolution[43]
  • Cryopreservation of sperm of Xenopus laevis and Xenopus tropicalis Michael G. Sargent, Timothy J. Mohun. 2005, Genesis [44]

Genotyping, Genome & DNA damage/repair

• • Sex determination in Xenopus laevis via RT-PCR. Amin Eimanifar, John Aufderheide, Suzanne Z. Schneider, Henry Krueger, and Sean Gallagher 2019. "Development of an in vitro diagnostic method to determine the genotypic sex of Xenopus laevis". PeerJ January 1, 2019; 7 e6886. [[45]]
  • Studying the DNA damage response in embryonic systems ElenaLo Furno, Bénédicte Recolin, Siemvan der Laan, Antoine Aze and Domenico Maiorano. 2021. Chapter Five in Methods in Enzymology Volume 661, 2021, Pages 95-120.[46]

Behavioral Assays

• • Precisely controlled visual stimulation to study experience-dependent neural plasticity in Xenopus tadpoles. Masaki Hiramoto and Hollis Cline. 2021. STAR Protocols | January 15, 2021. [[47]]

Cell cycle, Chromatin and Spindles

• • Immunochemical Detection of Modified Cytosine Species in Lampbrush Chromatin. Garry T. Morgan. 2021. In: Ruzov A., Gering M. (eds) DNA Modifications. Methods in Molecular Biology, vol 2198. Humana, New York, NY. [48]

Electrophysiology

• • K+ Release Assay and K+ Measurement in Oocyte Assay. Hong Li. 2020. Bio-Protocol 10(21),Nov5,2020, [DOI:10.21769/BioProtoc.3802]
  • A novel voltage-clamp/dye uptake assay reveals saturable transport of molecules through CALHM1 and connexin channels. Gaete et al 2021. J Gen Physiol. Nov 2; 152(11): e202012607. Published online 2020 Oct 19. doi: 10.1085/jgp.202012607. PMID:33074302.
  • Electrophysiological Approaches for the Study of Ion Channel Function Guiying Cui, Kirsten A. Cottrill and Nael A. McCarty. 2021. Structure and Function of Membrane Proteins pp 49-67. [49]

Xenopus oocyte cell free extract

• • DNA Damage Response in Xenopus laevis Cell-Free Extracts. Tomas Aparicio Casado and Jean Gautier 2021. In: Cell Cycle Checkpoints pp 103-144. Part of the Methods in Molecular Biology book series (MIMB, volume 2267). [50]
  • Microfluidic encapsulation of Xenopus laevis cell-free extracts using hydrogel photolithography. Zachary Geisterfer, John Oakey, Jesse Gatlin. 2020. STAR Protocols Volume 1, Issue 3, 18 December 2020, 100221 [[51]]
  • 'Affinity Purification of Label-free Tubulins from Xenopus Egg Extracts'. Sebastian Reusch, Abin Biswas, William Graham Hirst, SimoneReber. 2020. STAR Protocols Volume 1, Issue 3, 18 December 2020, 100151. [[52]]
  • Multi-purpose yolk-depleted embryo extracts.[a freon-free protocol] Veenstra GJ, Destrée OH, Wolffe AP. Translation of maternal TATA-binding protein mRNA potentiates basal but not activated transcription in Xenopus embryos at the midblastula transition. Mol Cell Biol. 1999 Dec;19(12):7972-82. doi: 10.1128/MCB.19.12.7972. PMID: 10567523; PMCID: PMC84882.

Xenopus oocyte expression system

• • The Xenopus Oocyte as an Expression System for Functional Analyses of Fish Aquaporins. François Chauvigné, Alba Ferré, Joan Cerdà. 2021. In: Dosch R. (eds) Germline Development in the Zebrafish. Methods in Molecular Biology, vol 2218. Humana, New York, NY. [[53]]

Gene knock out/knock down and Gene Editing

• • Vivo-Morpholinos: a non-peptide transporter delivers Morpholinos into a wide array of mouse tissues. Morcos et al 2008. Biotechniques December 1, 2008; 45 (6): 613-4, 616, 618 passim. [54] This technique has been used successfully in Xenopus, See Ivanova et al 2021. XB-ART-58553, PubMed ID: 34676214. [55]
  • Morpholino Studies in Xenopus Brain Development, Bestman and Cline. 2019, Brain Development pp 377-395, Part of the Methods in Molecular Biology book series (MIMB, volume 2047) [[56]]
  • Protocol for RNaseH-mediated RNA depletion, see Supplemental file from "Optimized design of antisense oligomers for targeted rRNA depletion" by Wesley A. Phelps, Anne E. Carlson, Miler T. Lee, 2020. Nucleic Acids Research, gkaa1072, [[57]]
  • Optimized design of antisense oligomers for targeted rRNA depletion. Wesley Phelps, Anne Carlson and Miler Lee. 2021. Nucleic Acids Res January 1, 2021; 49 (1): e5. [[58]] The Oligo-ASST Web tool is available at The Oligo-ASST Web tool is available at https://mtleelab.pitt.edu/oligo. Source code for the Web application and a command-line version of the program are available at https://github.com/MTLeeLab/oligo-asst.

RNA solubility, RNA interference (RNAi)

• • A fractionation-based protocol to investigate RNA solubility phase transition during Xenopus oocyte maturation. Hyojeong Hwang, Meng Ma, Jing Yang 2024. STAR protocols. https://doi.org/10.1016/j.xpro.2023.102830 (OA in Cell Press- download PDF here: https://www.sciencedirect.com/science/article/pii/S2666166723007979?via%3Dihub)

• • Application of the RNA interference technique to Xenopus embryos: Specific reduction of the β-catenin gene products by short double-stranded RNA produced by recombinant human Dicer Yuta Tagami, Takeshi Nishiyama, Michiko Omote, and Minoru Watanabe. 2021. Dev Growth Differ November 24, 2021;[59]

MassSpec & Proteomics

• • Quantitative Proteomics of Xenopus Embryos I, Sample Preparation. Meera Gupta, Matthew Sonnett, Lillia Ryazanova, Marc Presler, Martin Wühr . 2018. Methods Mol Biol 2018;1865:175-194. doi: 10.1007/978-1-4939-8784-9_13
  • High-throughput transporter identification for small molecules in cell factories with validation using Xenopus expression assays. See Transportome-wide engineering of Saccharomyces cerevisiae,. 2021. Wang et al. Metabolic Engineering.[60]
  • Uptake Assays in Xenopus laevis Oocytes Using Liquid Chromatography-mass Spectrometry to Detect Transport Activity Jorgenson et al 2021. bio-protocol. [61]
  • Synthetic hyperacetylation of nucleosomal histones (protocol showing Synthetic histone acetylation inhibits DNA replication). Kajino et al 2021. RSC Chemical Biology [62]
  • Xenopus laevis Oocytes Preparation for in-Cell EPR Spectroscopy (In-cell electron paramagnetic resonance (EPR) spectroscopy to determine the structure and dynamics of bio-macromolecules in the cell) Laura John and Malte Drescher. 2021. bio-protocol [63]

Cold Spring Harbor Xenopus Protocols Edited by Hazel Sive. published online 2017-2021. [64]

Note: External links to CSHL press are provided but a subscription fee is required for access most CSHL protocols.
All CSHL protocol articles are Copyright 2018, 2019, 2020 or 2021  Cold Spring Harbor Laboratory Press- Request permission to re-use images/figures accordingly.

Husbandry and Obtaining Eggs and Embryos

• • Raising and Maintaining Xenopus tropicalis from Tadpole to Adult. Maura Lane, Michael Slocum and Mustafa K Khokha. 2021 [65]
  • Obtaining Xenopus tropicalis Eggs. Maura Lane, Emily K Mis and Mustafa K Khokha 2021. [66]
  • Obtaining Xenopus tropicalis Embryos by Natural Mating. Maura Lane and and Mustafa K Khokha. 2021. [67]
  • Obtaining Xenopus tropicalis Embryos by In Vitro Fertilization. Maura Lane and and Mustafa K Khokha. 2021. [68]
  • Obtaining Xenopus laevis Eggs. Nikko Shaidani, Sean McNamara, Marcin Wlizla, Marko Horb. 2020. [69]
  • Obtaining Xenopus laevis Embryos. Nikko Shaidani, Sean McNamara, Marcin Wlizla, Marko Horb. 2020. [70]
  • Cryopreservation of Xenopus Sperm and In Vitro Fertilization Using Frozen Sperm Samples. Anna Nobel, Anita Abu-Daya, Matt Guille. 2021. [71]

General Research Protocols

• • Microinjection of Xenopus tropicalis Embryos. Maura Lane , Emily K Mis and Mustafa K Khokha. 2021. [72]
  • Whole-Mount In Situ Hybridization of Xenopus Embryos. Jean-Pierre Saint-Jeannet. 2017 [73]
  • Whole-Mount In Situ Hybridization of Xenopus Oocytes. Diana Bauermeister and Tomas Pieler. 2018. [74]
  • Fluorescence In Situ Hybridization of Cryosectioned Xenopus Oocytes. Christopher R. Neil and Kimberly Mowry. 2018. [75]
  • Whole-Mount Immunocytochemistry in Xenopus. Michael W. Klymkowsky. 2018. [76]
  • Microinjection of mRNAs and Oligonucleotides. Sally A. Moody. 2018. [77]
  • Microinjection of DNA Constructs into Xenopus Embryos for Gene Misexpression and cis-Regulatory Module Analysis. Yuuri Yasuoka and Masanori Taira. 2019. [78]
  • Whole-Mount Immunofluorescence for Visualizing Endogenous Protein and Injected RNA in Xenopus Oocytes. Samantha P. Jeschonek and Kimberly L. Mowry. 2018. [79]
  • Isolation of Xenopus Oocytes. Karen Newman, Tristan Aguero, and Mary Lou King. 2018. [80]
  • Microinjection of Xenopus Oocytes. Tristan Aguero, Karen Newman, and Mary Lou King. 2018. [81]
  • Oocyte Host-Transfer and Maternal mRNA Depletion Experiments in Xenopus. Douglas W. Houston. 2018. [82].
  • Applying Tensile and Compressive Force to Xenopus Animal Cap Tissue. Georgina K. Goddard, Nawseen Tarannum, and Sarah Woolner. 2020. [83].
  • RNAi-Mediated Loss of Function of Xenopus Immune Genes by Transgenesis. Eva-Stina Edholm and Jacques Robert. 2018. [84]
  • Flow Cytometric Analysis of Xenopus Immune Cells. Eva-Stina Edholm. [85]
  • Isolation and Demembranation of Xenopus Sperm Nuclei. James W. Hazel and Jesse C. Gatlin. 2018. [86]
  • Patch-Clamp and Perfusion Techniques to Study Ion Channels Expressed in Xenopus Oocytes. Guohui Zhang and Jianmin Cui. 2018. [87]
  • Heterologous Protein Expression in the Xenopus Oocyte. Jonathan S. Marchant. 2018. [88]
  • Isolation and Analysis of Xenopus Germinal Vesicles. Garry T. Morgan. 2018. [89]
  • Xenopus Tadpole Tissue Harvest. Matthew D. Patmann, Leena H. Shewade, Katelin A. Schneider, and Daniel R. Buchholz. 2017 [90].
  • In Vivo Transfection of Naked DNA into Xenopus Tadpole Tail Muscle. Lindsey Marshall, Fabrice Girardot, Barbara A. Demeneix, and Laurent Coen. 2017. [91]
  • Cell Proliferation Analysis during Xenopus Metamorphosis: Using 5-Ethynyl-2-Deoxyuridine (EdU) to Stain Proliferating Intestinal Cells. Morihiro Okada and Yun-Bo Shi. 2017. [92]

Genome Editing Protocols

• • Generating Nonmosaic Mutants in Xenopus Using CRISPR-Cas in Oocytes Sang-Wook Cha. 2021 [93]
  • CRISPR-Cas9 Mutagenesis in Xenopus tropicalis for Phenotypic Analyses in the F0 Generation and Beyond.Ira Blitz and Takuya Nakayama. 2021 [94]
  • Modeling Human Genetic Disorders with CRISPR Technologies in Xenopus. helen Willsey, Matt Guille, and Robert Grainger. 2021. [95]

'Omics

• • Transcriptomics and Proteomics Methods for Xenopus Embryos and Tissues. Michael J. Gilchrist, Gert Jan C. Veenstra, and Ken W.Y. Cho. 2020. ''Topic Introduction'' [96]
  • Mass Spectrometry-Based Absolute Quantification of Single Xenopus Embryo Proteomes. Rik G.H. Lindeboom, Arne H. Smits, Matteo Perino, Gert Jan C. Veenstra, and Michiel Vermeulen. [97]
  • INTACT Proteomics in Xenopus. Lauren Wasson, Nirav M. Amin, and Frank L. Conlon. 2019. [98]

Imaging

• • Xenopus Tadpole Craniocardiac Imaging Using Optical Coherence Tomography. Engin Deniz, Emily K Mis, Maura Lane and Mustafa K Khokha. 2021. [99]
  • Live Imaging of Cytoskeletal Dynamics in Embryonic Xenopus laevis Growth Cones and Neural Crest Cells. Burcu Erdogan, Elizabeth Bearce, Laura Anne Lowery. 2020. [100]
  • Imaging Methods in "Xenopus" Cells, Embryos, and Tadpoles. Lance A Davidson and Laura Anne Lowery. 2021. [101]
  • Imaging Structural and Functional Dynamics in Xenopus Neurons. Hollis Cline. 2021 [102]

Genomes, Chromosomes, DNA, Chromatin and Epigenetics

• • In Vitro Transcription Systems. Michael R. Green and Joseph Sambrook. [103]
  • ChIP-Sequencing in Xenopus Embryos. Saartje Hontelez, Ila van Kruijsbergen, and Gert Jan C. Veenstra. 2019. [104]
  • Chromatin Characterization in Xenopus laevis Cell-Free Egg Extracts and Embryos. Wei-Lin Wang, Takashi Onikubo, and David Shechter. 2019. [105].
  • Analysis of Chromatin Binding of Ectopically Expressed Proteins in Early Xenopus Embryos. Laura J.A. Hardwick and Anna Philpott. 2019. [106].
  • Analysis of Phosphorylation Status of Ectopically Expressed Proteins in Early Xenopus Embryos. Laura J.A. Hardwick and Anna Philpott. 2019. [107].
  • Assessing Ubiquitylation of Individual Proteins Using Xenopus Extract Systems. Gary S. McDowell and Anna Philpott. 2019. [108]
  • Generating a Three-Dimensional Genome from Xenopus with Hi-C. Ian K. Quigley and Sven Heinz. 2019. [109]
  • An RNA-Seq Protocol for Differential Expression Analysis. Nick D.L. Owens, Elena De Domenico, and Michael J. Gilchrist. 2019. [110]
  • DNase-seq to Study Chromatin Accessibility in Early Xenopus tropicalis Embryos. Jin Sun Cho, Ira L. Blitz, and Ken W.Y. Cho. 2019 [111]
  • Mapping Chromatin Features of Xenopus Embryos. George E. Gentsch and James C. Smith. 2019. [112]
  • Reconstituting Nuclear and Chromosome Dynamics Using Xenopus Extracts. Susannah Rankin. 2019 [113]
  • Extracts for Analysis of DNA Replication in a Nucleus-Free System. Justin Sparks and Johannes C. Walter. 2019. [114]
  • Endoplasmic Reticulum Network Formation with Xenopus Egg Extracts. Songyu Wang, Fabian B. Romano, and Tom A. Rapoport. 2019. [115].
  • Chromosome Cohesion and Condensation in Xenopus Egg Extracts. Eulália M.L. da Silva and Susannah Rankin. 2019. [116]
  • Chromatin Interaction Analysis Using Paired-End-Tag (ChIA-PET) Sequencing in Tadpole Tissues. Nicolas Buisine, Xiaoan Ruan, Yijun Ruan, and Laurent M. Sachs. 2018. [117]
  • Chromatin Immunoprecipitation for Chromatin Interaction Analysis Using Paired-End-Tag (ChIA-PET) Sequencing in Tadpole Tissues. 2018. Nicolas Buisine, Xiaoan Ruan, Yijun Ruan, and Laurent M. Sachs. [118]

Endocrinology, Toxicology and Metamorphosis

• • Methods for Investigating the Larval Period and Metamorphosis in Xenopus. Daniel R. Buchholz and Yun-Bo Shi. ''Topic Introduction'' [119].
  • Frog Embryo Teratogenesis Assay—Xenopus (FETAX): Use in Alternative Preclinical Safety Assessment. Douglas J. Fort and Michael Mathis. 2018. [120]
  • Following Endocrine-Disrupting Effects on Gene Expression in Xenopus laevis. Petra Spirhanzlova, Michelle Leemans, Barbara A. Demeneix, and Jean-Baptiste Fini. 2019. Full-text link: [121].
  • Larval Thymectomy of Xenopus laevis. Sara Mashoof, Breanna Breaux, and Michael F. Criscitiello. 2018. [122]

Cell-free extracts, Cell-free systems

• • The Use of Cell-Free Xenopus Extracts to Investigate Cytoplasmic Events. Romain Gibeaux and Rebecca Heald. 2019.''Topic Introduction'' [123]
  • Calculating the Degradation Rate of Individual Proteins Using Xenopus Extract Systems. Gary S. McDowell and Anna Philpott. 2019. [124].
  • Special Considerations for Making Explants and Transplants with Xenopus tropicalis. 2019. Marilyn Fisher and Robert M. Grainger. [125]. Supplemental Material [126].
  • Filopodia-Like Structure Formation from Xenopus Egg Extracts. Helen M. Fox and Jennifer L. Gallop. 2019. [127].
  • Centromere and Kinetochore Assembly in Xenopus laevis Egg Extract. Julio C. Flores Servin and Aaron F. Straight. 2018. [128]
  • Dissecting Protein Complexes in Branching Microtubule Nucleation Using Meiotic Xenopus Egg Extracts. Jae-Geun Song and Sabine Petry. 2018. [129]
  • Protein Immunodepletion and Complementation in Xenopus laevis Egg Extracts. Christopher Jenness, David J. Wynne, and Hironori Funabiki. 2018. [130]
  • Analysis of Mitotic Checkpoint Function in Xenopus Egg Extracts. Yinghui Mao. 2018 [131]
  • Chemical Screening Using Cell-Free Xenopus Egg Extract. Matthew R. Broadus and Ethan Lee. 2018. [132]
  • Robustly Cycling Xenopus laevis Cell-Free Extracts in Teflon Chambers. Jeremy B. Chang and James E. Ferrell Jr. [133]
  • Microfluidic Encapsulation of Demembranated Sperm Nuclei in Xenopus Egg Extracts. John Oakey and Jesse C. Gatlin. 2018. [134]
  • Nucleus Assembly and Import in Xenopus laevis Egg Extract. Pan Chen and Daniel L. Levy. 2018. [135]
  • Assembly of Spindles and Asters in Xenopus Egg Extracts. Christine M. Field and Timothy J. Mitchison. 2018. [136]
  • Preparation of Cellular Extracts from Xenopus Eggs and Embryos. Matthew C. Good and Rebecca Heald. 2018. [137]
  • Using the Xenopus Oocyte Toolbox. Kim L. Mowry. 2020. [138]
  • The Use of Xenopus for Cell Biology Applications. Anna Philpott. 2020. [139]

Immunobiology

• • Cryosectioning and Immunostaining of Xenopus Embryonic Tissues. Olga Ossipova and Sergei Y Sokol. 2021. [140]
  • Raising Antibodies for Use in Xenopus. Maya Z. Piccinni and Matthew J. Guille. 2020. [141]
  • Purifying Antibodies Raised against Xenopus Peptides. Maya Z. Piccinni and Matthew J. Guille. 2020. [142]
  • Assessing the Immune Response When Raising Antibodies for Use in Xenopus. Maya Z. Piccinni and Matthew J. Guille. 2020. [143].
  • Lymphocyte Deficiency Induced by Sublethal Irradiation in Xenopus. Louise A. Rollins-Smith and Jacques Robert. 2019. [144].
  • Skin Grafting in Xenopus laevis: A Technique for Assessing Development and Immunological Disparity. Yumi Izutsu. 2019. [145]
  • Adoptive Transfer of Fluorescently Labeled Immune Cells in Xenopus. Kun Hyoe Rhoo and Jacques Robert. [146]
  • Assessing Antibody Responses to Pathogens or Model Antigens in Xenopus by Enzyme-Linked Immunosorbent Assay (ELISA). Francisco De Jesús Andino and Jacques Robert. 2019 [147]
  • In Vivo Assessment of Neural Precursor Cell Cycle Kinetics in the Amphibian Retina. Morgane Locker and Muriel Perron2019. [148], Supplemental Material [149]
  • Protein Immunodepletion and Complementation in Xenopus laevis Egg Extracts. Christopher Jenness, David J. Wynne, and Hironori Funabiki. 2018. [150]
  • Elicitation of Xenopus laevis Tadpole and Adult Frog Peritoneal Leukocytes. Leon Grayfer. 2018. [151]
  • RNAi-Mediated Loss of Function of Xenopus Immune Genes by Transgenesis. Eva-Stina Edholm and Jacques Robert. 2018. [152]
  • Flow Cytometric Analysis of Xenopus Immune Cells. Eva-Stina Edholm. [153]

Assessing Behaviour and Behavioural Phenotypes

• • Schooling in Xenopus laevis Tadpoles as a Way to Assess Their Neural Development. Virgillio Lopez III, Arseny Khakhalin and CArlos Aizenman. 2021. [154]

Neurobiology, Electrophysiology and Tissue Regeneration

• • Tetrode Recording in the Xenopus laevis Visual System Using Multichannel Glass Electrodes.M Hiramoto and Hollis Cline. 2021 [155]
  • Electrophysiological Approaches to Studying Normal and Abnormal Retinotectal Circuit Development in the Xenopus Tadpole. Kara Pratt. 2021 [156]
  • Making In Situ Whole-Cell Patch-Clamp Recordings from Xenopus laevis Tadpole Neurons. WC Li. 2021. [157]
  • Analysis of Visual Collision Avoidance in Xenopus Tadpoles. Arseny Khakhalin. 2020 [158]
  • Ex Vivo Eye Tissue Culture Methods for Xenopus. Jonathan J. Henry, Kimberly J. Perry, and Paul W. Hamilton. 2019. [159].
  • Methods for Examining Lens Regeneration in Xenopus. Jonathan J. Henry, Kimberly J. Perry, and Paul W. Hamilton. 2019. [160]
  • Tracing Central Nervous System Axon Regeneration in Xenopus. Kurt M. Gibbs and Ben G. Szaro. 2018. [161]
  • Cell Transplantation as a Method to Investigate Spinal Cord Regeneration in Regenerative and Nonregenerative Xenopus Stages. Emilio E. Méndez-Olivos and Juan Larraín. 2018. [162]
  • Infrared Laser-Mediated Gene Induction at the Single-Cell Level in the Regenerating Tail of Xenopus laevis Tadpoles. Riho Hasugata, Shinichi Hayashi, Aiko Kawasumi-Kita, Joe Sakamoto, Yasuhiro Kamei, and Hitoshi Yokoyama. 2018. [163]
  • Rod-Specific Ablation Using the Nitroreductase/Metronidazole System to Investigate Regeneration in Xenopus. Reyna I. Martinez-De Luna and Michael E. Zuber. 2018. [164]
  • Studies of Limb Regeneration in Larval Xenopus. Anthony L. Mescher and Anton W. Neff. 2019. [165].
  • Bulk Electroporation-Mediated Gene Transfer into Xenopus Tadpole Brain. Cristina Sáenz de Miera, Ethan Parr, and Robert J. Denver. 2018. [166] . Supplemental Material [167].
  • Inverse Drug Screening of Bioelectric Signaling and Neurotransmitter Roles: Illustrated Using a Xenopus Tail Regeneration Assay. Kelly G. Sullivan and Michael Levin. 2018. [168]
  • Neurophysiological and Behavioral Analysis in Xenopus. Ben Szaro. 2021. (Electrophysiology protocols for studies of neural circuitry in the developing optic tectum and spinal cord, vocalizations, and locomotory behavior).

[169]

• • Electrophysiological Recording for Study of Xenopus Retinotectal Circuitry. Y Luo , W Shen, and Hollis Cline. 2021. [170]
  • Bulk Dye Loading for In Vivo Calcium Imaging of Visual Responses in Populations of Xenopus Tectal Neurons. PW Hogg and K Haas. 2021. [171]
  • Xenopus, a Model to Study Wound Healing and Regeneration: Experimental Approaches. Paula Slater, M Palacios, Juan Larraín. 2021. [172]
  • In Vivo Time-Lapse Imaging and Analysis of Dendritic Structural Plasticity in Xenopus laevis Tadpoles. Hai-Yun He, Chih-Yang Lin and Hollis Cline. 2021. [doi: 10.1101/pdb.prot106781] [173]

Fate mapping, Explants and Transplants

• • Dissecting and Culturing Animal Cap Explants. Kevin S. Dingwell and James C. Smith. 2018. [174]
  • Cranial Neural Crest Explants. Hélène Cousin and Dominique Alfandari. 2018. [175]
  • Einsteck Transplants. Hélène Cousin. 2019 [176]
  • Cleavage Blastomere Deletion and Transplantation to Test Cell Fate Commitment in Xenopus. Sally A. Moody. 2019. [177]
  • Cleavage Blastomere Explant Culture in Xenopus. Sally A. Moody. 2019. [178]
  • Lineage Tracing and Fate Mapping in Xenopus Embryos. Sally A. Moody. 2018. [179]
  • Collagen-Embedded Tumor Transplantations in Xenopus laevis Tadpoles. Maureen Banach and Jacques Robert. 2017 [180]
  • Organ Culture of the Xenopus Tadpole Intestine. Atsuko Ishizuya-Oka. 2017 [181]

CSHL Recipes

• • Embryo Lysis Buffer (Xenopus). (Recipe 1) CSHLP. 2019 [182]
  • Xenopus Embryo Lysis Buffer. (Recipe 2). CSHLP. 2019 [183]
  • Lysis Buffer for Xenopus (recipe3) CSHLP. 2019. [184]
  • DNA Isolation Buffer. CSHLP. 2019 [185]
  • Lysis Buffer for Xenopus Hi-C. CSHLP 2019. [186]
  • RIPA Buffer for Xenopus. CSHLP 2019. [187]
  • Amphibian Serum-Free (ASF) Medium Supplemented with Fetal Bovine Serum (FBS). CSHL 2019. [188]
  • Xenopus Eye Culture Medium. CSHLP 2019. [189]
  • Marc's Modified Ringer's (MMR) for Xenopus (1×). CSHLP. 2018. [190]
  • Marc's Modified Ringer's (MMR) for Xenopus (20×). CSHLP. 2019. [191]
  • Injection Buffer for Xenopus. CSHLP 2019. [192]
  • TE Buffer for Xenopus. CSHLP 2019. [193]
  • Proteinase K Buffer for Xenopus Oocytes. CSHLP 2018. [194]
  • PBT/PBT-Plus for Xenopus Oocytes. CSHL. 2018. [195]
  • Energy Mix for Xenopus Egg Extracts. CSHL 2018. [196]
  • Xenopus Oocyte Culture Medium (XOCM).CSHL 2018.[197]
  • PBT for Xenopus Oocyte FISH.CSHL 2018. [198]
  • Blocking Solution for Xenopus Oocytes. CSHL 2018. [199]
  • Aldehyde Fixative (MEMFA). CSHL 2018. [200]
  • Staining Buffer for Xenopus Embryos. CSHL 2017. [201]
  • Blocking Solution for Xenopus Embryos. CSHL 2017. [202]
  • Embryonic Xenopus Culture Media (CM). CSHL 2017. [203]
  • Hybridization Buffer (HB) for Xenopus Embryos. CSHL 2017. [204]

Cold Spring Harbor Xenopus Protocols 2007. Edited by Hazel Sive

• • Housing and Feeding of Xenopus laevis - Sive et al. [205]
  • Inducing Ovulation in Xenopus laevis - Sive et al. [206]
  • Xenopus laevis In Vitro Fertilization and Natural Mating Methods - Sive et al. [207]
  • Egg Collection and In Vitro Fertilization of the Western Clawed Frog Xenopus tropicalis - Showell & Conlon [208]
  • Isolation of Xenopus Oocytes - Sive et al. [209]
  • Isolating Xenopus laevis Testes - Sive et al. [210]
  • Dejellying Xenopus laevis Embryos - Sive et al. [211]
  • Removing the Vitelline Membrane from Xenopus laevis Embryos - Sive et al. [212]
  • Microinjection of Xenopus Embryos - Sive et al. [213]
  • Defolliculation of Xenopus Oocytes - Sive et al. [214]
  • Microinjection of Xenopus Oocytes - Sive et al. [215]
  • Animal Cap Isolation from Xenopus laevis - Sive et al. [216]
  • Xenopus laevis Keller Explants - Sive et al. [217]
  • Microinjection of RNA and Preparation of Secreted Proteins from Xenopus Oocytes - Sive et al. [218]
  • Calibration of the Injection Volume for Microinjection of Xenopus Oocytes and Embryos - Sive et al. [219]
  • Isolation of DNA from Red Blood Cells in Xenopus - Sive et al. [220]
  • Investigating Morphogenesis in Xenopus Embryos: Imaging Strategies, Processing, and Analysis - Kim & Davidson [221]
  • Low-Magnification Live Imaging of Xenopus Embryos for Cell and Developmental Biology - Wallingford [222]
  • High-Magnification In Vivo Imaging of Xenopus Embryos for Cell and Developmental Biology - Keiserman et al. [223]
  • Preparation of Fixed Xenopus Embryos for Confocal Imaging - Wallingford [224]
  • Whole-Mount Fluorescence Immunocytochemistry on Xenopus Embryos - Lee et al. [225]
  • Generation of Transgenic Xenopus laevis - Kroll & Amaya - [226] [227] [228]
  • In Vivo Time-Lapse Imaging of Neuronal Development in Xenopus - Ruthazer et al. [229]
  • Photoconversion for Tracking the Dynamics of Cell Movement in Xenopus laevis Embryos - Chernet et al. [230]
  • Single-Cell Electroporation in Xenopus - Liu & Haas [231]
  • Imaging Axon Pathfinding in Xenopus In Vivo - Leung & Holt [232]
  • A Versatile Protocol for mRNA Electroporation of Xenopus laevis Embryos - Chernet & Levin [233]

General Research Protocols

Animal Husbandry

• obtain frogs and frog housing equipment [234]
• Xenopus Express links [235] - Xenopus Express husbandry [236]
• Xenopus tropicalis and Xenopus laevis environmental parameter standard operating procedure (Kroll lab)

Lab Solutions and Reagents (click each to view expanded content)

• 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)

Transgenesis

• 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). [237]
• A method for generating transgenic frog embryos. Ishibashi S, Kroll KL, Amaya E. Methods Mol Biol. 2008;461:447-66. [238]
• A method for generating transgenic frog embryos. Amaya E, Kroll KL. Methods Mol Biol. 1999;97:393-414. [239]
• Optimization Strategies for the CRISPR–Cas9 Genome-Editing System. Vejnar et al 2016. [240]

in situ Hybridization

• in situ hybridization, brief version (Harland lab)
• whole-mount in situ hybridization (Conlon lab)
  • in situ checklist (Conlon lab)
• whole-mount in situ hybridization - LNA probes (Wheeler lab)
• fluorescent in situs and FCIS (Vize lab)
  • Wholemount embryo FISH / Double FISH
  • Flourescin Tyramide Synthesis
  • Cy3 Tyramide Synthesis
  • FCIS
• RNA Whole-Mount In situ Hybridisation Proximity Ligation Assay (rISH-PLA) (Myers and Scarlett labs)

Immunohistochemistry

• 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)
• RNA Whole-Mount In situ Hybridisation Proximity Ligation Assay (rISH-PLA) (Myers and Scarlett labs)

ChIP protocols

• Chromatin immunoprecipitation analysis of Xenopus embryos., Methods Mol Biol. 2012;917:279-92. [241] [242]
  • Akkers RC, Jacobi UG, Veenstra GJ.
• Chromatin immunoprecipitation in early Xenopus laevis embryos., Dev Dyn. 2009 Jun;238(6):1422-32. [243] [244]
  • Blythe SA, Reid CD, Kessler DS, Klein PS.

Histology

• cryosectioning ''Xenopus'' embryos (Conlon lab)

Embryo Staining Protocols (non in situ)

• 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)

Immuno 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)
• RNaseH-mediated RNA depletion
  • Optimized design of antisense oligomers for targeted rRNA depletion (Phelps et al 2020)) [[245]]

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)
• frozen egg extracts that retain spindle assembly activity (Takagi and Shimamoto, 2017)

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)