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ABSTRACT ARCHIVE |
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CryoLetters is a bimonthly, international journal for low temperature science and technology |
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CryoLetters 31 (6), 438-444 (2010) A SIMPLE OSMOTIC STRESS TEST TO PREDICT BOAR SPERM CRYOSURVIVAL Cesar Garzon-Perez1, Hector F. Flores2 and Alfredo Medrano1* 1Departamento de Ciencias Pecuarias, Facultad de Estudios Superiores – Cuautitlan, Universidad Nacional Autonoma de Mexico, Carretera Cuautitlan –Teoloyucan s/n. Cuautitlan
Izcalli, Estado de Mexico. 54714. Mexico Abstract This work was carried out to test whether viability of pig spermatozoa subjected to an osmotic test is correlated to sperm cryosurvival. Spermatozoa were cooled from 22°C to -5°C, aliquots were exposed to a series of hyperosmotic solutions (300-2100 mOsm/kg) for 15 min, immediately spermatozoa were re-warmed to 37ºC and isosmolarity was restored. Spermatozoa were cooled from 22°C to -5°C and one aliquot was exposed to the osmotic test while diluted spermatozoa were frozen-thawed. Plasma membrane-intact spermatozoa decreased as osmolarity increased (P<0.0001), a further decreased (P<0.0001) was observed when isotonicity was restored. Proportions of plasma membrane-intact and acrosome-intact cells from the osmotic test were no different from those after freeze-thawing: 36% vs. 35%, 80% vs. 80%, respectively. A significant correlation was found between the proportion of acrosome-intact cells after freeze-thawing and that from the osmotic test (r=0.81, P<0.01). This test provides a useful and economical mean to predict in vitro boar sperm cryosurvival. Keywords: semen cooling, chlortetracycline assay, sperm capacitation, lectins
CryoLetters 31 (6), 445-453 (2010) SUPERCOOLING CAPACITY AND COLD HARDINESS OF THE ADULTS OF THE SYCAMORE LACE BUG, Corythucha ciliata (HEMIPTERA: Tingidae Rui-Ting Ju1, 2*, Feng Wang2, Yu-Yu Xiao2 and Bo Li1** 1Coastal Ecosystems Research Station of the Yangtze River Estuary, Ministry of Education Key Laboratory for Biodiversity Science and Ecological Engineering, Institute of Biodiversity
Science, Fudan University, Shanghai 200433, P. R. China. Abstract Supercooling point (SCP) of female adults of Corythucha ciliata was significantly lower than that of male adults, with an average being -11.49°C and -9.54°C, respectively. Low temperature survival of adults of different ages indicated that there were differences in cold survival ability among age groups of adults. Nonlinear regression analysis found that the response of C. ciliata adults to exposure time under different low temperature regimes (above -5°C) was best fitted by a logistic equation. Both low temperature and exposure time had significant effects on mortality of adults. Temperatures above 5°C did not prevent C. ciliata adults from surviving. C. ciliata was shown to be a freeze-intolerant but chill-tolerant insect. C. ciliata could tolerate subzero temperatures by supercooling. Temperature around -8°C is a critical point for successful overwintering of C. ciliata adults, which can establish in the whole areas where Platanus trees are planted in China. Keywords: cold hardiness, Corythucha ciliata, sycamore lace bug, supercooling, Platanus
CryoLetters 31 (6), 454-459 (2010) IN VITRO AND IN VIVO SURVIVAL OF MOUSE BLASTOCYSTS AFTER REPEATED VITRIFICATION WITH THE OPEN PULLED STRAW (OPS) METHOD M. El-Gayar 1, 2, M. Gauly 1 and W. Holtz 1* 1Department of Animal Science, Georg-August-University, Albrecht-Thaer-Weg 3, 37075 Goettingen, Germany. Abstract This study addresses the in vitro and in vivo survival of mouse embryos repeatedly vitrified by the OPS-technique of Vajta et al. (24). Of 225 vitrified blastocysts, after warming one third was cultured in vitro, the other two-thirds were re-vitrified. After these were warmed, the second third was put to culture. With the remaining third the procedure was repeated. Of embryos vitrified once, 97% developed to expanded blastocysts and 81% proceeded to hatching. Corresponding values for embryos re-vitrified once were 93% and 72%, respectively (P>0.05). After another re-vitrification, expansion and hatching rates were reduced to 76% and 35%, respectively (P<0.01). Of 10 recipients provided with 10 embryos each that had been vitrified once, 80% remained pregnant with 5.5 ± 0.3 fetuses. Corresponding values for re-vitrified embryos were 80% and 5.0 ± 0.3. Of all embryos transferred, 44% became vital fetuses after a single vitrification, compared to 40% after re-vitrification. Keywords: Mouse, Cryopreservation, Repeated vitrification, Open pulled straw, Embryo transfer.
CryoLetters 31 (6), 460-472 (2010) ENCAPSULATION DEHYDRATION COLLIGATIVE CRYOPROTECTIVE STRATEGIES AND AMPLIFIED FRAGMENT LENGTH POLYMORPHISM MARKERS TO VERIFY THE IDENTITY AND GENETIC STABILITY OF Euglenoids FOLLOWING CRYOPRESERVATION Keith Harding1*, Julia Müller2, Hella Timmermann2, Maike Lorenz2, John G. Day3 and Thomas Friedl2 1Plant Conservation Group, University of Abertay Dundee (*current address: Damar Research Scientists, Damar, Drum Road, Cuparmuir, Fife, KY15 5RJ, UK) Abstract An encapsulation/dehydration procedure was developed for Euglena gracilis Klebs as a ‘model alga’ to examine various cryoprotective regimes combined with controlled rate cooling to cryopreserve other Euglenoid taxa. Cryoprotective variables were optimised to enable reproducible growth following a combination of alginate encapsulation, sucrose osmotic dehydration, air desiccation, methanol treatment, cooling to -40°C and plunging into liquid nitrogen (LN). Amplified Fragment Length Polymorphism (AFLP) analysis was adapted to: (i) verify algal identity by discriminating between different Euglenoids and (ii) examine the genetic stability of algal cultures prior to various stages of cryoprotective treatments and following exposure to LN. AFLPs were highly reproducible (>99%) as reliable diagnostic markers, where a single DNA fragment change accounted for ~0.4% of the detectable variation in an AFLP pattern. AFLP changes were detected in cryoprotective treatments following LN exposure. Successive stages of the dehydration and desiccation treatments did not accumulate AFLP changes indicating these are random events. Keywords: AFLP; cryopreservation; encapsulation dehydration; genetic stability; microalgae
CryoLetters 31 (6), 473-484 (2010) CRYOPRESERVATION OF HAIRY ROOTS OF Rubia akane (NAKAI) USING A DROPLET-VITRIFICATION PROCEDURE Haeng-Hoon Kim1* , Elena V. Popova1, Jung-Yoon Yi1, Gyu-Taek Cho1, Sang-Un Park2, Sheong-Chun Lee3 and Florent Engelmann4, 5 1 National Agrobiodiversity Center, National Academy of Agricultural Science, RDA, Suwon 441-707, Korea. 2 Division of Plant Science and Resources, Chungnam National University,
Daejeon 305-764, Korea. Abstract An efficient protocol for the cryopreservation of madder (Rubia akane Nakai) hairy root cultures was developed using droplet-vitrification and alternative loading and vitrification solutions formulated previously in our laboratory. Among eight preculture treatments tested, the highest post-cryopreservation regeneration was obtained for explants incubated in liquid half-strength MS medium with progressively increased sucrose concentration (0.3 M for 54 h, then 0.5 M for 16 h). Loading of precultured explants improved their post-cryopreservation regeneration by 50-75% compared with non-loaded control. Combining loading solution C4 (35% PVS3) and vitrification solution B5 (80% PVS3) was the most effective, while applying six PVS2-based solutions at room temperature resulted in low post-cryo regeneration. Treatment duration was optimized to 30 min for loading and to 10-20 min for vitrification solution. Apices of primary and secondary hairy roots showed similar post-cryo regeneration (88 and 95%, respectively), which was significantly higher than regeneration of root sections without apices (65%). Droplet-vitrification produced higher post-cryo regeneration than ‘classical’ vitrification in cryovials. Our results suggest that droplet-vitrification using alternative loading and vitrification solutions is an efficient method for cryopreservation of R. akane hairy root cultures. Keywords: droplet-vitrification, hairy roots, loading, preculture, Rubia akane, vitrification solution.
CryoLetters 31 (6), 485-492 (2010) CRYOPRESERVATION OF CHICK ISLETS Datar, S.P.1 and Bhonde, R.R.2* 1 S. P. College, Tilak Road, Pune-411030, India. email: savitapdatar@yahoo.co.in Abstract The avian endocrine pancreas shares some similarities with those of mammals. Previously we have reported a technique of isolation of B islets from chick pancreas and also demonstrated their possible use for hypoglycemic drug screening as efficiently as mammalian islets. Here we describe a novel cryopreservative medium for the cryopreservation of chick islets. Isolated chick islets were suspended in a cryo medium consisting of Dulbecco’s modified Minimum Essential Medium: Ham’s F12 (1:1), Fetal Bovine Serum (20%), Dimethylsulfoxide (10%) with different concentrations (50µg/ml to 500 µg/ml) of riboflavin or nicotinamide. The viability of revived islets after three and half months was checked by trypan blue dye exclusion and functionality was assessed by insulin secretion in response to glucose challenge. The maximum recovery of viable islets and insulin secretion was obtained in response to glucose challenge at 250µg/ml concentration of Riboflavin or Nicotinamide. This is a first report on cryopreservation of chick islets exhibiting cryoprotective role of water soluble vitamins without vitamin C. Keywords: Chick islets, Cryopreservation, Riboflavin, Nicotinamide.
CryoLetters 31 (6), 493-503 (2010) DIFFUSION OF DIMETHYL SULFOXIDE IN TISSUE ENGINEERED COLLAGEN SCAFFOLDS VISUALIZED BY COMPUTER TOMOGRAPHY Inga Bernemann1*, Navid Manuchehrabadi1,2, Ralf Spindler1, Jeunghwan Choi2, Wim F. Wolkers1, John C. Bischof2# and Birgit Glasmacher1# 1Institute for Multiphase Processes, Leibniz Universitaet Hannover, Germany, Abstract Cryopreservation is a convenient method for long-term preservation of natural and engineered tissues in regenerative medicine. Homogeneous loading of tissues with CPAs, however, forms one of the major hurdles in tissue cryopreservation. In this study, computer tomography (CT) as a non-invasive imaging method was used to determine the effective diffusion of Me2SO in tissue-engineered collagen scaffolds. The dimensions of the scaffolds were 30 x 30 x 10mm3 with a homogeneous pore size of 100 µm and a porosity of 98%. CT images were acquired after equilibrating the scaffolds in phosphate buffered saline (PBS) and transferring them directly in 10% (v/v) Me2SO. The Me2SO loading process of the scaffold could thus be measured and visualized in real time. The experimental data were fitted using a diffusion equation. The calculated effective diffusion constant for Me2SO in the PBS loaded scaffold was determined from experimental diffusion studies to be 2.4 x 10-6 cm2/s at 20°C. Keywords: Computer Tomography, Visualization, Chemical Loading, Freezing of Tissue Engineered Products, Collagen Scaffolds, CPA Diffusion.
CryoLetters 31 (6), 504-512 (2010) RAPID COLD-HARDENING IN Zaprionus vittiger (COQUILLETT) (DIPTERA: Drosophilidae) Casper Nyamukondiwa* and John S. Terblanche Department of Conservation Ecology and Entomology, Faculty of AgriSciences, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa ABSTRACT Low temperature injury can be significantly reduced by pre-treatment at sub-lethal low temperatures of only a few hours -a phenomenon termed ‘rapid cold-hardening’ (RCH). However, most studies to date have focused extensively on only a few key insect taxa, e.g. Drosophila melanogaster within the Drosophilidae family. Further studies on other closely-related species are required to better understand evolutionary and ecological variation in the magnitude of the RCH response in terrestrial arthropods. Here, we investigated RCH in a previously unstudied fruit fly, Zaprionus vittiger, following a range of high and low pretreatment temperatures. There was a significant improvement in Z. vittiger survival of lethal temperatures (2 h at -3°C) following a 2 h pretreatment at 4, 7 and 10°C as well as 30°C for 2 h. Maximum survival (60-70%) during RCH was achieved following hardening at 7°C and 10°C but is lower than some Drosophila species under similar treatment conditions. Therefore, since RCH was detected in a confamilial species, we propose that RCH might be a widely conserved response to temperature variation in the family Drosophilidae, although some variation in the magnitude of the response can be detected. Keywords: Thermal history, cold tolerance, rapid cold-hardening, phenotypic plasticity.
CryoLetters 31 (6), 513-524 (2010) FACETED ICE GROWTH IN HYDROPHILIC AND HYDROPHOBIC CHANNELS Y.C. Liu, C.H. Tsai and C.W. Lan* Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan 10617, R.O.C. Abstract This study is motivated by the cryo-preservation for tissues, which is related to the solidification of blood in a confined space. The morphological behavior of growing ice facet in 1.3 mm hydrophilic and hydrophobic channels is the focus of this study. Both DI water and KCl solution were solidified without seeds in the channels and the growing front facet was discussed in regards to contact angle, in-situ morphology, conductivity difference between two phases and solute segregation. For DI water, the formation of facets was observed during ice growth in both hydrophilic and hydrophobic channels and could be classified into two modes according to the local curvature of the interface. The facet was formed at different positions of the interface in hydrophilic and hydrophobic channels. Solutal segregation effect was taken into consideration to partially govern the interface morphology and the facet formation during solidification of the KCl solution. In addition, the growth rates of the ice growth with a facet were also compared and discussed. Keywords: facet, ice growth, hydrophilic, hydrophobic, channel |
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For Abstracts published from meetings, such as SLTB meetings, go to the
relevant Volume Year of the journal (above). For Full text Free Access Content (from 2000 onwards) go to CryoLetters at Ingenta and look for the blue symbol. |
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