Characterization of kidney marrow in zebrafish (Danio rerio) by using a new surgical technique.

Zebrafish kidney marrow (ZKM), which is equivalent to the haematopoietic bone marrow of mammals, produces all major blood cell types, which morphologically resemble their mammalian counterparts. To be able to exploit the advantages of zebrafish genetics for analysis of the general mechanisms controlling self-renewal, proliferation and lineage decisions of vertebrate haematopoetic cell populations, it is essential to develop a simple surgical technique in order to identify, dissect and take out the ZKM without contamination with other surrounding tissues and cells. However, the size of adult zebrafish is small (average size: 2.5 cm) and the ZKM is an extremely protected organ and not easy to localize, which makes this procedure a great microsurgical challenge. Here we report a new microsurgical technique to identify, localize and dissect ZKM in adult zebrafish using a new approach. The potential advantages of this technique are summarized here: it allows purity of the sample, which is critical for performing flow cytometry analysis and/or cell number count; it enables visualization of the ZKM without a parenchimal incision, which simplifies the further dissection; the learning curve is short, requiring only basic microsurgical skills, and it is reliable and highly reproducible. To further characterize the kidney marrow cells obtained by this technique, we performed histology, flow cytometry, cytospin experiments and cell counts.