Biospin spectrometer

To synthesize methacrylated dextran with various degrees of functionalities, dextran (2.0 g, Mw: 40–500 kDa) was dissolved in 10 mL of anhydrous dimethyl sulfoxide (DMSO) with the addition to 0.2 g of base catalyst 4-dimethylamino pyridine (DMAP) and the required molar equivalent of glycidyl methacrylate (GMA, density = 1.042 g/mL at 25 °C). The mixture solution was kept constant at 45 °C and stirred for 24 h. After stirring, the reaction solution was pipetted dropwise into a 200 mL of ice-chilled isopropanol to precipitate modified dextran. The precipitation was then collected via centrifugation and subsequently re-dissolved and dialyzed against 4 L of milli-Q water at a temperature of 4 °C preceding lyophilization. Purified methacrylated-dextran (Dex-MA) was stored at − 20 °C until use. The functionality and degree of methacrylation of Dex-MA were analyzed using nuclear magnetic resonance spectroscopy (NMR, a 700 MHz Bruker BioSpin spectrometer, in D₂O/DMSO), where the degree of methacrylation confirmed to achieve 40–88% modification depending on the initial molar equivalent of glycidyl methacrylate to glucopyranose. The peaks at 5.75 and 6.2 ppm represent the protons at the double bond of the GMA group.

B. subtilis colonies (3610 and Δmatrix) were grown on LB supplemented with ¹³C-labeled glucose for 72 h to provide a ¹³C-enrichment and were filled in 4 mm SSNMR rotors. Experiments were performed on a 600 MHz Bruker Biospin spectrometer equipped with a 4 mm Efree probe at a magic-angle spinning frequency of 11 kHz. Cross-polarization spectra were recorded using 1024 scans and a contact time of 1 ms. Water-edited ¹³C spectra (256 scans) were recorded using a T₂-filter of 1.5 ms followed by a variable ¹H mixing time. The 2D ¹H-¹³C HETCOR experiments were recorded with and without a 5 ms ¹H-¹H mixing time using 128 scans and acquisition times of 7.5 ms (indirect) and 20 ms (direct).

Detailed approaches for recovering explanted human hearts and functional and structural imaging of the atria were described previously [6 (link),12 (link)]. Intact explanted human donor hearts (n = 7, 47 ± 14 y.o., two females) with AF history and/or cardiac co-morbidities were obtained from Lifeline of Ohio Organ Procurement Organization (table 1). The Institutional Review Board defined the study on samples from deceased donors as Not Human Subjects Research. The entire atria were isolated, coronary perfused, immobilized with blebbistatin and stained with the voltage-sensitive dye di-4ANDBQBS for near-infrared optical mapping (UltimaL CMOS camera, SciMedia, Japan) [4 (link),6 (link)]. Induction of AF by burst pacing (2–10 s) from the superior RA and/or posterior LA was attempted in all hearts. AF drivers were defined as localized sites with the most repetitive and recurrent re-entrant pattern that led to neighbouring regions and had a minimum temporal stability of 30%. After the functional mapping, CE-MRI was performed using a 9.4 T Bruker BioSpin spectrometer (Ettlingen, Germany) [4 (link),6 (link)].