Quantifying the Growth Rate and Morphology of Ice Crystals Growth in Cryoprotectants Via High-Speed Camera and Cryomicroscope

Shu

2017

Sci Rep

Gas-liquid-solid phase transition behaviour of water is studied with environmental scanning electron microscopy for the first time. Abnormal phenomena are observed. At a fixed pressure of 450 Pa, with the temperature set to −7 °C, direct desublimation happens, and ice grows continuously along the substrate surface. At 550 Pa, although ice is the stable phase according to the phase diagram, metastable liquid droplets first nucleate and grow to ~100–200 μm sizes. Ice crystals nucleate within the large sized droplets, grow up and fill up the droplets. Later, the ice crystals grow continuously through desublimation. At 600 Pa, the metastable liquid grows quickly, with some ice nuclei floating in it, and the liquid-solid coexistence state exists for a long time. By lowering the vapour pressure and/or increasing the substrate temperature, ice sublimates into vapour phase, and especially, the remaining ice forms a porous structure due to preferential sublimation in the concave regions, which can be explained with surface tension effect. Interestingly, although it should be forbidden for ice to transform into liquid phase when the temperature is well below 0 °C, liquid like droplets form during the ice sublimation process, which is attributed to the surface tension effect and the quasiliquid layers.

mentioning

confidence: 99%

Abnormal gas-liquid-solid phase transition behaviour of water observed with in situ environmental SEM

Shu

2017

Sci Rep

Journal of Heat Transfer

“…It should be noted, though, that these temperature gradients also exist in convectively warmed and cooled droplets, so the relative magnitude of the gradients is important when considering error in the measurements. Future studies may incorporate more advanced image processing techniques to quantitatively determine the amount of ice in droplets systems, such as those used in the measurement of ice growth rates in thin films of CPAs [ 40 ].…”

Section: Resultsmentioning

confidence: 99%

Ultra-Rapid Laser Calorimetry for the Assessment of Crystallization in Low-Concentration Cryoprotectants

Zhan

Liu

Natesan

et al. 2022

Cryoprotective agents (CPAs) are routinely used to vitrify, attain an amorphous glass state void of crystallization, and thereby cryopreserve biomaterials. Two vital characteristics of a CPA loaded system are the critical cooling and warming rates (CCR and CWR), the temperature rates needed to achieve and return from a vitrified state respectively. Due to the toxicity associated with CPAs, it is often desirable to use the lowest concentrations possible, driving up CWR and making it increasingly difficult to measure. This paper describes a novel method for assessing CWR between the 0.4×105-107 °C/min in microliter CPA loaded droplet systems with a new ultra-rapid laser calorimetric approach. Cooling was achieved by direct quenching in liquid nitrogen, while warming was achieved by the irradiation of plasmonic gold nanoparticle-loaded vitrified droplets by a high-power 1064 nm millisecond pulsed laser. We assume "apparent" vitrification is achieved provided ice is not visually apparent (i.e. opacity) upon imaging with a camera during cooling or highspeed camera during warming. Using this approach, we were able to investigate CWR in single CPA systems such as glycerol, PG, and Trehalose in water, and mixtures of glycerol-trehalose-water and propylene glycol-trehalose-water CPA at low concentrations (20-40 wt%). Further, an phenomenological model for determining the CCRs and CWRs of CPA was developed which allowed for predictions of CCR or CWR of single component CPA and mixtures, providing an avenue for optimizing CCR and CWR and perhaps future CPA cocktail discovery.

“…Then upon rapid cooling to −60 °C with a cooling rate of 40 °C min –1 , a homogeneous solution instantly produced numerous tiny ice crystal grains, and the temperature was carefully raised to allow most of the ice polycrystals to melt until only one single ice crystal was left, and the growth behavior of a single ice crystal at the supercooling temperature (Δ T 1 ) of 0.5 °C was recorded. As illustrated in Figures b and S3, the radius of the ice crystals (i.e., the radius of curvature at the specific points on the curve was obtained through fitting the curve of the edge of ice crystals by the polynomial function) in 5.0% EG and 5.0% EG + 20 mg mL –1 Ficoll 70 (Fic, a positive control, a widely used commercial CPA) were 346.4 ± 82.7 and 189.6 ± 32.6 μm at 50 s, respectively, which was reduced by 243 μm with the addition of 20 mg mL –1 SF (103.6 ± 32.6 μm), suggesting the capability of SF on inhibiting the ice growth in 5.0% EG is better than that of Fic. Moreover, the curvature of the single ice crystal in 5.0% EG was 0.15 μm –1 , which decreased significantly to 0.08 μm –1 with the addition of SF, indicating the smoother boundaries of ice crystals recrystallized in SF solution (Figure S4).…”

Section: Results and Discussionmentioning

confidence: 99%

“…Prior to quickly cooling the cryostage to −60 °C. The images were processed using Canny Edge Extraction Algorithm to calculate the ice curvature …”

Section: Experimental Sectionmentioning

confidence: 99%

See 1 more Smart Citation

Strong Hydration Ability of Silk Fibroin Suppresses Formation and Recrystallization of Ice Crystals During Cryopreservation

et al. 2021

The cryopreservation (CP) of cell/tissue is indispensable in medical science. However, the formation of ice during cooling and ice recrystallization/growth in time of thawing present significant risk of cell/tissue damage upon analysis of CP process. Herein, the natural and biocompatible silk fibroin (SF) with regular hydrophobic and hydrophilic domains, were first employed as a cryoprotectant (CPA), to the CP of human bone-derived mesenchymal stem cells (hBMSCs), which has been routinely cyropreserved for cell-based therapies. Addtion of SF can regulate the formation of ice crystals during cooling process because of its strong hydration ability in the comparation to the cryopreservation medium (CM) without SF. Moreover, the devitrification-induced recrystallization/growth of ice during the thawing process is suppressed. Most importantly, the addition of 10 mg mL–1 SF can achieve 81.28% cell viability of cryopreserved hBMSCs as similar as those with the addition of 180 mg mL–1 Ficoll 70 (commercial CPA), and the functions of the cryopreserved hBMSCs are maintained as good as that of the fresh ones. This work is not only significant for meeting the ever-increasing demand of cell therapy, but also trailblazing for designing materials in controlling ice formation and growth during the CP of other cells and tissues.