Phosphate buffered saline pbs

The three species of deep‐sea shrimps used in this study, Parasergestes armatus, Allosergestes sargassi, and Deosergestes henseni, represent the three major organs of Pesta morphologies (i.e., bilobed, trilobed, and fringed, respectively; Figure 1b–d). The animals for this study were obtained during two deep‐sea research expeditions, the first occurring in the Straits of Florida on May 4–8, 2019, on the R/V Weatherbird and the second occurring in the Northern Gulf of Mexico on June 9–22, 2019, as part of a NOAA Ocean Research Exploration expedition on the R/V Point Sur. Animals were captured by 1 or 9‐m² tucker trawl over sampling events that occurred both day and night over a total range of depths from 150–1,500 m. Animals captured on the first expedition were immediately placed in 4% paraformaldehyde in 0.1 M phosphate‐buffered saline (PBS; Boston BioProducts, Ashland, MA, USA) for a 48‐hr fixation step prior to transfer into 0.1 M PBS for subsequent analyses. Those from the second expedition were examined immediately, allowing us to obtain morphological data from fresh animals prior to fixation.

Secreted antibodies were purified by Protein A chromatography. Harvested culture supernatants were first flocculated with polyDADMAC (EMD Millipore, Burlington, MA), followed by centrifugation for 20 minutes at 15,000 g. The clarified supernatants were filtered and applied to 5 mL Praesto Jetted A50 resin (Purolite, King of Prussia, PA) in a XK26/20 column (Cytiva, Marlborough, MA) using an AKTA explorer FPLC. The column was first equilibrated with phosphate-buffered saline (PBS) (Boston BioProducts, Ashland, MA). Prior to elution with 0.1 M acetic acid, the column was washed with 10 column volumes PBS. The pH of the eluted fractions was adjusted by adding 1 M Tris-HCl (pH 9).