Isolation and Maintenance of In Vitro Cell Cultures from the Ctenophore Mnemiopsis leidyi

Abstract

The ability to isolate, monitor, and examine specific cells of interest enables targeted experimental manipulations that would otherwise be difficult to perform and interpret in the context of the whole organism. In vitro primary cell cultures derived from ctenophores thus serve as an important tool for understanding complex cellular and molecular interactions that take place both within and between various ctenophore cell types. Here we describe methods for reliably generating and maintaining primary cell cultures derived from the lobate ctenophore Mnemiopsis leidyi that can be used for a wide variety of experimental applications.

Extracellular DNA traps in a ctenophore demonstrate immune cell behaviors in a non-bilaterian

“…To investigate whether the model ctenophore Mnemiopsis leidyi possesses cell types capable of specialized immune functions, we mechanically disassociated whole Mnemiopsis and examined isolated cells 47 , 48 . We observed morphologically distinct motile cell types displaying prominent intracellular granules and/or vesicles, including amoebocyte-like cells with relatively short pseudopodia and stellate cells with long pseudopodia (Fig.…”

“…Laboratory cultures of Mnemiopsis leidyi were maintained as previously described 44 . Adult Mnemiopsis cells were isolated from whole animals following established protocols 32 , 47 , 48 , 77 . In brief, whole small adult Mnemiopsis (approximately 3 cm in length) were homogenized to a single-cell preparation using a loose-fitting dounce homogenizer.…”

Extracellular DNA traps in a ctenophore demonstrate conserved immune cell behaviors in a non-bilaterian

“…To investigate whether the model ctenophore Mnemiopsis leidyi possesses cell types with specialized immune functions, we mechanically disassociated whole Mnemiopsis and examined isolated cells 46,47 . We observed morphologically distinct motile cell types displaying prominent intracellular granules and/or vesicles, including amoebocyte-like cells with relatively short pseudopodia and stellate cells with long pseudopodia ( Fig.…”

“…Laboratory cultures of Mnemiopsis leidyi were maintained as previously described (Presnell, 2021 #35). Adult Mnemiopsis cells were isolated from whole animals following established protocols 36,46,47 . Crassostrea gigas were maintained under flowing seawater at approximately 13°C and hemolymph was extracted from the adductor muscle with a syringe.…”

“…Live cell staining was performed following (2) and live imaging was performed using a JuLI Stage (NanoEntek). For immunofluorescence, ctenophore cells were prepared as previously described 46,47,73 and labeled with mouse anti-histone H11-4 (EMD Millipore) and anti-mouse Alexa Fluor 488 (Thermo Fisher Scientific). The H11-4 histone antibody was selected because it recognizes histones H1, H2A/B, H3, and H4 proteins across diverse species.…”