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

Methods in Molecular Biology
Ryan J Debuque, James W Godwin

Abstract

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.

Citations

Apr 3, 2021·Developmental Dynamics : an Official Publication of the American Association of Anatomists·Ryan J DebuqueJames W Godwin

❮ Previous
Next ❯

Related Concepts

Related Feeds

Antibody Repertoire Diversity

Antibody repertoire diversity and its role during natural infection is a prerequisite for molecular and structural elucidation of functionally protective immunity. Discover the latest insights into antibody diversity here.

Related Papers

Methods in Molecular Biology
Louise Edvardsson, Tor Olofsson
Current Protocols in Stem Cell Biology
Antonija Kreso, Catherine Adell O'Brien
Journal of Immunological Methods
Alexander R PintoJames W Godwin
Methods in Molecular Biology
Tetsutaro Hayashi, Kiyokazu Agata
© 2022 Meta ULC. All rights reserved