Cameron Scheithauer's research works | University of Minnesota Duluth and other places

Fig. 1: Schematic flow of steps (left to right) in liter scale vitrification and rewarming. Liter volumes of a CPA (0.5-3L) in cryobags are large enough to hold a human organ. The cryobag is placed inside a controlled rate freezer (CRF) for cooling. For nanowarming (top section of the

Fig. 2: Demonstration of physical success of vitrification in multiple volumes. a Table summarizes vitrification results for all the 3 CPAs and volumes. Photos of a successful vitrified (glass) M22 inside a cryobag for b 0.5 Liter, c 1 Liter, and d 3 Liter (largest volume reported). The out-of-plane thicknesses are 5.5, 6.5, and 10.5 cm for 0.5, 1, and 3L cryobags, respectively.

$Fig. 3: Thermal results from experimental and modeled liter scale CPA vitrification. a Schematic for a representative case, 0.5 L cryobag containing CPA with placement of three fiber optic temperature probes (3 cm apart). Blue arrows show the direction of LN2 flow in CRF. b Experimental and predicted temperature vs. time plot for 0.5L M22. The dashed green line shows the programmed CRF temperature profile/protocol. c CRF cooling protocols for 0.5, 1, and 3 L volumes. The regions of ice formation and fracture danger are labeled. Scatter plot of d center cooling rate and e temperature difference (ΔTmax in the glassy region) for all three volumes tested for M22 (mean ± SD; n=3). Cooling rate is calculated in ranges 0 to -100 °C and -120 to -150 °C for temperature difference plots. Mean cooling rates are greater than the CCR of M22 (~0.1°C/min). Temperature differences are within the allowable limit (dashed) (< 20°C) calculated from a simple thermal shock equation [20].$

Fig. 4: Photos of the porcine liver (left) before (T = 4°C) and (right) after vitrification (T = -150°C). The pattern in the photo was due to the cryobag placement on a supporting mesh in the control rate freezer (CRF) (see Fig. S7B). The cryobag was removed for the vitrified liver photo to reduce glare and get a clear photo.

Fig. 6: Nanowarming specific absorption rate (SAR). a Plot of SARFe (SARV/ CFe) vs. magnetic field strength (H) measured at room temperature for iron-oxide nanoparticles IONPs (sIONPs in M22 shown here) at two frequencies (190 and 360kHz) (plotted mean ± SD; n=3). b Plot of SARFe vs. temperature for sIONPs in M22. Average SARFe (mean ± SD; n=3) is plotted in three different temperature regions, i.e., glass, supercooled, and liquid. SAR is measured from

Physical vitrification and nanowarming at human organ scale to enable cryopreservation

November 2024

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Organ banking by vitrification could revolutionize transplant medicine. However, vitrification and rewarming have never been demonstrated at the human organ scale. Using modeling and experimentation, we tested the ability to vitrify and rewarm 0.5–3 L volumes of three common cryoprotective agent (CPA) solutions: M22, VS55, and 40% EG+0.6M Sucrose. We first demonstrated our ability to avoid ice formation by convectively cooling faster than the critical cooling rates of these CPAs while also maintaining adequate uniformity to avoid cracking. Vitrification success was then verified by visual, thermometry, and x-ray μCT inspection. M22 and EG+sucrose were successfully vitrified in 0.5 L bags, but only M22 was vitrified at 3 L. VS55 did not vitrify at any tested volumes. As additional proof of principle, we successfully vitrified a porcine liver (~1L) after perfusion loading with 40% EG+0.6M Sucrose. Uniform volumetric rewarming was then achieved in up to 2 L volumes (M22 with ~5 mgFe/mL iron-oxide nanoparticles) using nanowarming, reaching a rate of ~88 °C/min with a newly developed 120 kW radiofrequency (RF) coil operating at 35kA/m and 360kHz. This work demonstrates that human organ scale vitrification and rewarming is physically achievable, thereby contributing to technology that enables human organ banking.

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Cameron Scheithauer’s research while affiliated with University of Minnesota Duluth and other places

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