PUBLICATION
Establishment of novel monoclonal antibodies for identification of type A spermatogonia in teleosts1
- Authors
- Hayashi, M., Ichida, K., Sadaie, S., Miwa, M., Fujihara, R., Nagasaka, Y., Yoshizaki, G.
- ID
- ZDB-PUB-190512-7
- Date
- 2019
- Source
- Biology of reproduction 101(2): 478-491 (Journal)
- Registered Authors
- Yoshizaki, Goro
- Keywords
- fish, germ cell transplantation, monoclonal antibodies, rainbow trout, spermatogonia, zebrafish
- MeSH Terms
-
- Animals
- Animals, Genetically Modified
- Antibodies, Monoclonal/immunology*
- Breeding
- Enzyme-Linked Immunosorbent Assay
- Epitopes
- Female
- Male
- Mice
- Mice, Inbred BALB C
- Oncorhynchus mykiss/physiology*
- Spermatogenesis/genetics
- Spermatogenesis/physiology
- Spermatogonia/classification
- Spermatogonia/immunology
- Spermatogonia/physiology*
- PubMed
- 31077286 Full text @ Biol. Reprod.
Citation
Hayashi, M., Ichida, K., Sadaie, S., Miwa, M., Fujihara, R., Nagasaka, Y., Yoshizaki, G. (2019) Establishment of novel monoclonal antibodies for identification of type A spermatogonia in teleosts1. Biology of reproduction. 101(2):478-491.
Abstract
We recently established a germ cell transplantation system in salmonids. Donor germ cells transplanted into the body cavity of recipient embryos migrate toward and are incorporated into the recipient gonad, where they undergo gametogenesis. Among the various types of testicular germ cells, only type A spermatogonia (A-SG) can be incorporated into the recipient gonads. Enriching for A-SG is therefore important for improving the efficiency of germ cell transplantation. To enrich for A-SG, an antibody against a cell surface marker is a convenient and powerful approach used in mammals; however, little is known about cell surface markers for A-SG in fish. To that end, we have produced novel monoclonal antibodies (mAbs) against cell-surface molecules of rainbow trout (Oncorhynchus mykiss) A-SG. We inoculated mice with living A-SG isolated from pvasa-GFP transgenic rainbow trout using GFP-dependent flow cytometry. By fusing lymph node cells of the inoculated mice with myeloma cells, we generated 576 hybridomas. To identify hybridomas that produce mAbs capable of labeling A-SG preferentially and effectively, we screened them using cell ELISA, fluorescence microscopy, and flow cytometry. We thereby identified two mAbs that can label A-SG. By using flow cytometry with these two antibodies, we could enrich for A-SG with transplantability to recipient gonads from amongst total testicular cells. Furthermore, one of these mAbs could also label zebrafish (Danio rerio) spermatogonia. Thus, we expect these monoclonal antibodies to be powerful tools for germ cell biology and biotechnology.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping