Methods for axolotl blood collection, intravenous injection, and efficient leukocyte isolation from peripheral blood and the regenerating limb

Ryan Debuque, James Walter Godwin

Research output: Chapter in Book/Report/Conference proceedingChapter (Book)Otherpeer-review

2 Citations (Scopus)


The vertebrate immune system comprises both adaptive and innate immune cells with distinct functions during the resolution of inflammation and wound healing after tissue injury. Recent evidence implicates a requirement for innate immune cells from the myeloid lineage during the early stages of limb regeneration in the Mexican axolotl. Understanding the functions of innate and adaptive immune cells in the axolotl has been hampered by a lack of approaches to isolate and analyze these cells. Here we describe a protocol to isolate myeloid cells from the regenerating axolotl limb that incorporates intravenous delivery of physiological labels. In addition we provide a protocol to enrich for leukocytes in the peripheral blood. These protocols produce single-cell suspensions that can be analyzed using flow cytometry or sorted into specific subsets using fluorescent-activated cell sorting (FACS). FACS is a routine approach to sort cells based on their physical characteristics as well as their cell surface antigen repertoire. Isolated cell populations can then be analyzed in a wide range of downstream assays to facilitate a greater understanding of leukocyte biology in the axolotl.
Original languageEnglish
Title of host publicationSalamanders in Regeneration Reseach
Subtitle of host publicationMethods and Protocols
EditorsAnoop Kumar, Aandras Simon
Place of PublicationTotowa NJ USA
PublisherHumana Press
Number of pages22
ISBN (Print)9781493924943
Publication statusPublished - 2015

Publication series

NameMethods in Molecular Biology

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