Dataset: Carbohydrate microarray (epitope) analyses of POM-derived carbohydrates in the Western North Atlantic Ocean in May 2019

Collection

Water was collected via Niskin bottles mounted on a rosette, equipped with a CTD in the western North Atlantic Ocean aboard R/V Endeavor (EN638) during May 2019. Seawater was transferred to carboys that were rinsed three times with water from the sampling depth and then filled with seawater from a single Niskin bottle.

Particulate organic matter (POM) was harvested by filtering between 5-15 liters of seawater through a 47-mm pre-combusted (400℃ for 6 hours) glass fiber filter (GF/F; nominal pore size 0.7 μm; for volumes filtered at each depth and station, see the dataset). Filers were stored at -80C until further analysis.  The same filter was used for both particulate organic carbon (POC) measurements and monosaccharide composition analysis.   

Analysis

Polysaccharide extraction for microarray analyses of POM

POM samples were prepared for polysaccharide analysis according to Vidal-Melgosa et al. (2021). Polysaccharides were sequentially extracted from four filter piece punches (11.2 mm diameter) from GF/F filters. The samples were first extracted with autoclaved MilliQ water, followed by 50 mM EDTA, and finally 4 M NaOH with 0.1% NaBH₄. The supernatant containing extracted polysaccharides was collected from each of the sequential steps and stored at 4 °C.

Carbohydrate microarray analysis to determine structural complexity of POM

The polysaccharides extracted as described above were analyzed following Vidal-Melgosa et al. (2021). In brief, the polysaccharide extracts were first diluted in printing buffer (55.2% glycerol, 44% water, 0.8% Triton X-100), and then printed on 0.45 µm pore size nitrocellulose membrane (Whatman) using a microarray robot (Sprint, Arrayjet, Roslin, UK) at 20 °C and 50% humidity. The membranes were probed with one of 9 monoclonal antibodies, washed multiple times, and probed with secondary antibodies (anti-rat, anti-mouse, or anti-His tag) conjugated to alkaline phosphatase for 2 hours. The arrays were developed using 5-bromo-4-chloro-3-indolyphosphate and nitro blue tetrazolium in alkaline phosphatase buffer (100 mM NaCl, 5 mM MgCl₂, 100 mM Tris-HCl, pH 9.5). The microarrays were scanned and signal intensity was acquired using the software Array-Pro Analyzer 6.3 (Media Cybernetics). Signals were normalized among samples; higher signals correspond to a higher abundance of a given polysaccharide epitope. Note that the carbohydrate microarray data are only semiquantitative; while comparisons can be made for the abundance of a given epitope between stations and depths, the signal intensity cannot be used to compare signals of different epitopes.