Schock, E. | Search bioRxiv

bioRxiv2022

Production and Characterization of Monoclonal antibodies to Xenopus Proteins

Monoclonal antibodies are powerful and versatile tools that enable the study of proteins in diverse contexts. They are often utilized to assist with identifying subcellular localization and characterizing the function of target proteins of interest. However, because there can be considerable sequence diversity between orthologous proteins in Xenopus and mammals, antibodies produced against mouse or human proteins often do not recognize Xenopus counterparts. To address this issue, we refined existing protocols to produce mouse monoclonal antibodies directed against Xenopus proteins of interest. Here we describe several approaches for the generation of useful mouse anti-Xenopus antibodies to multiple Xenopus proteins and their validation in various experimental approaches. These novel antibodies are now available to the research community through the Developmental Study Hybridoma Bank (DSHB). Summary statementThe manuscript describes the generation and characterization of novel monoclonal antibodies to Xenopus laevis proteins using refined hybridoma production methods suitable for basic science research labs.

developmental biology↗

bioRxiv2018

A transition from SoxB1 to SoxE transcription factors is essential for progression from pluripotent blastula cells to neural crest cells

The neural crest is a stem cell population unique to vertebrate embryos that gives rise to derivatives from multiple embryonic germ layers. The molecular underpinnings of potency that govern neural crest potential is highly conserved with that of pluripotent blastula stem cells, suggesting that neural crest cells may have evolved through retention of aspects of the pluripotency gene regulatory network (GRN). A striking difference in the regulatory factors utilized in pluripotent blastula cells and neural crest cells is the deployment of different subfamilies of Sox transcription factors; SoxB1 factors play central roles in the pluripotency of naive blastula and ES cells, whereas neural crest cells require SoxE function. Here we explore the shared and distinct activities of these factors to shed light on the role that this molecular hand-off of Sox factor activity plays in the genesis of neural crest and the lineages derived from it. Our findings provide evidence that SoxB1 and SoxE factors have both overlapping and distinct activities in regulating pluripotency and lineage restriction in the embryo. We hypothesize that SoxE factors may transiently replace SoxB1 factors to control pluripotency in neural crest cells, and then poise these cells to contribute to glial, chondrogenic and melanocyte lineages at stages when SoxB1 factors promote neuronal progenitor formation.

developmental biology↗