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Abstracts: CryoLetters 26 (5), 2005

CryoLetters 26 (5), 277-278 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

EDITORIAL:
OUR STATED ETHICAL POLICY

 

 

CryoLetters 26 (5), 279-288 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

THE INFLUENCE OF 1-10 kD FRACTION FROM BRAINS OF THE HIBERNATING GROUND SQUIRREL AND THE YAKUT HORSE ON PROLIFERATION AND PROTEIN SYNTHESIZING SYSTEM OF EHRLICH ASCITIC CARCINOMA CELLS

A.K. Gulevsky*, V.I. Grischenko, O.S. Tereschenko and I.J. Shchenyavcky

Institute for Problems of Cryobiology and Cryomedicine of the National Akademy of Sciences of Ukraine, 23 Pereyaslavskaya. Street, 61015 Kharkov-15, Ukraine.
*Corresponding author:
ivanou@rambler.ru

Abstract

The experimental data presented in the work testify to the cytostatic activity of 1-10 kD polypeptide fractions from brains of the hibernating ground squirrel and the Yakut horse towards Ehrlich ascitic carcinoma (EAC) cells. The experiments on the investigation of the inhibiting influence of 1-10 kD fractions from tissues of the hibernating and cold-adapted animals on protein-synthesizing system of EAC cells allow us to conclude that the cytostatic effect of the fractions is effected at the genetic level in the tumor cells.

Keywords: cold-adapted animals, polypeptide fractions, cytostatic activity.

 

 

CryoLetters 26 (5), 289-296 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

THERMAL ANALYSIS OF TERTIARY BUTYL ALCOHOL/SUCROSE/WATER TERNARY SYSTEM

Jian-Guo Zuo, Tse-Chao Hua*, Bao-Lin Liu and Guo-Yan Zhou

Institute of Cryomedicine and Food Freezing, Shanghai University of Science and Technology, No.516 Jun-Gong Road, Shanghai 200093, China. 
*Corresponding author: Tel.: 86-21-65685291. Fax: 86-21-65685291
E-mail:
tchua@sh163.net

Abstract

The purpose of this work is to investigate the freezing properties of tertiary butyl alcohol (TBA)/sucrose/water ternary system. Differential scanning calorimetry (DSC) is employed to determine the glass transition temperature of the maximally freeze-concentrated solution Tg' and the crystallization (or devitrification) temperature Tr. DSC measurements show that the presence of sucrose hinders the crystallization of TBA during cooling. The residual TBA in the glassy state will cause a decrease in Tg' and will crystallize during heating. An increase in the cooling rate causes a decrease in Tg'. For 10% TBA/10% sucrose/water ternary system, the critical heating rate is approximately 250C/min. Annealing treatment at temperatures below Tg' causes the crystallization of TBA, which indicates that TBA molecules still have appreciable mobility even at temperatures below Tg'. When the ratio of TBA to sucrose is less than 0.2, TBA cannot crystallize during cooling.

Keywords: freeze-drying, tertiary butyl alcohol (TBA), sucrose, glass transition, differential scanning calorimetry (DSC), annealing

 

 

CryoLetters 26 (5), 297-304 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

RUMINATIONS ON HEAT AND MASS TRANSFER IN FREEZE DRYING: A PERSONAL REFLECTION

Terence Rowe

Castro Urdiales, Balmaseda, Spain. Contact: PMatejt@nibsc.ac.uk

Abstract

Examples are given to clarify the difference between heat and temperature, the sequence involving the passage of heat to and within a sample undergoing freeze-drying at reduced pressure is scrutinised and the beneficial and counterproductive effects of "modifying the vacuum" are examined together with the mechanism involved. The commentator also glances at possibly undesirable contamination of the product and the system as well as questioning the various forms of measurement of vacuum and temperature and their significance in validation procedures. An assertion is made that if the process is correctly conducted. The sublimation phase will be a square law function of the product depth and not a linear one. A proposal is made that heat input should be based on a variable energy profile (with appropriate safeguards) rather than on the basis of shelf temperature.

 

 

CryoLetters 26 (5), 305-312 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

EFFECT OF COOLING AND EXPOSURE TO ETHYLENE GLYCOL ON IN VITRO MATURATION AND EMBRYO DEVELOPMENT OF RHESUS OOCYTES

Catherine A. VandeVoort 1, 2, * and S. P. Leibo 3, 4

1California Regional Primate Research Center;
2Department of Obstetrics and Gynecology, School of Medicine, University of California, Davis, CA, USA.
3Department of Biological Sciences, University of New Orleans;
4Audubon Center for Research of Endangered Species; New Orleans, Louisiana, USA.

Abstract

The meiotic spindle of metaphase II-stage oocytes is damaged when mature oocytes are cooled to temperatures close to 0C, as occurs during cryopreservation by equilibrium cooling. Since a spindle has not yet formed within a germinal vesicle-stage oocyte, it has been suggested that immature oocytes may be more resistant than metaphase II oocytes to cryopreservation by equilibrium cooling. To test this proposition, we examined the effects on rhesus macaque oocytes of chilling and exposure to ethylene glycol (EG) on their maturation and embryo development. A total of 202 cumulusintact oocytes was collected from adult female rhesus monkeys that had been given follicle stimulating hormone for controlled ovarian hyperstimulation. Within two hours of their having been aspirated and prior to germinal vesicle breakdown, oocytes were either cooled to 0C for 10 minutes or were exposed for 15 minutes at 35C to 1.5 M EG to be tested as a possible cryoprotectant. After being exposed, oocytes were cultured in maturation medium, fertilized in vitro with rhesus spermatozoa, and cultured. The maturation rate and subsequent development into blastocysts of those oocytes that had been exposed to EG or cooled to 0C did not differ significantly from untreated control oocytes. Additional germinal vesicle oocytes were exposed to 1.5 M EG at 35C for 3 minutes and then supercooled to 7C or frozen at 7C or frozen at 0.5C to 35C. Rates of maturation and embryo development of oocytes cooled to or frozen at 7C were significantly lower than rates for control oocytes; none of those frozen to 35C even underwent maturation. These results suggest that germinal vesiclestage oocytes may be less susceptible to injury resulting from chilling or exposure to ethylene glycol, but are still damaged by freezing.

Keywords: Rhesus macaque, cryopreservation, oocytes, cooling, ethylene glycol.

 

 

CryoLetters 26 (5), 313-322 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

PLANT REGROWTH FROM POTATO SHOOT TIPS CRYOPRESERVED BY A COMBINED VITRIFICATION-DROPLET METHOD

A. Halmagyi*, C. Deliu and A. Coste

Institute of Biological Research, 48 Republicii str. RO-400015, Cluj-Napoca, Romania.
*Corresponding author: e.mail:
halmagyi.a@gmx.net

Abstract

 Shoot tips obtained from in vitro potato plants (three cultivars) were successfully cryopreserved by the combined vitrification-droplet method and subsequently regenerated shoots. The effect of apex size, sucrose concentration, preculture duration and cold hardening treatments on viability of cryopreserved shoot tips was studied. The excised shoot tips were incubated, precultured and dehydrated with concentrated PVS2 cryoprotective solution at room temperature, prior to a direct plunge in liquid nitrogen. After rapid rewarming in Murashige-Skoog (MS) liquid medium, shoot tips were plated on semisolid MS medium (3.5 g/l agar) supplemented with 0.4 mg l-1 gibberellic acid, 0.5 mg l-1 zeatin, 0.2 mg l-1 indole-3-acetic acid and 30 g l-1 sucrose for regrowth. Cryopreserved shoot tips resumed growth within 20 days and regenerated shoots within 30 days. The highest regrowth levels of apices after cryopreservation were 55% recovery for cv. Dsire, 51% for cv. Ostara and 46% for cv. Sant.

Keywords: vitrification, shoot tips, axillary buds, preculture, dehydration.

 

 

CryoLetters 26 (5), 323-332 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

CRYOPRESERVATION OF APPLE USING NON-DESICCATED SECTIONS FROM WINTER-COLLECTED SCIONS

Leigh E. Towill and Remi Bonnart

USDA-ARS National Center for Genetic Resources Preservation, Fort Collins, CO 80521

Abstract

Winter vegetative buds of Malus species are cryopreserved at USDA-ARS NCGRP to backup genetic resources maintained by field collections. The method uses desiccation of nodal sections prior to cooling but is time and labor intensive, and can damage materials if excessive. Here we tested cooling sections without prior desiccation to improve the efficiency of handling. Sections were slowly cooled to -30C or -35C and transferred to the vapor phase over liquid nitrogen (LNV). Viability was assessed using a sprouting test or grafting test. Some accessions showed higher viability when cooled at 5C/day compared to 1C/h and when transferred to LNV at -35C compared to -30C. Ten of 20 species had accessions that were successfully cryopreserved using a criterion of 50% or greater sprouting.  Desiccation prior to cooling was not necessary for cryopreservation of winter-collected scions from these Malus species.

Keywords: conservation, germplasm, Malus species, two-step cooling, grafting.

 

 

CryoLetters 26 (5), 333-340 (2005)
© CryoLetters, c/o Royal Veterinary College, London NW1 0TU, UK

ULTRASTRUCTURAL IMPLICATIONS OF PRETREATMENT FOR SUCCESSFUL CRYOPRESERVATION OF ONCIDIUM PROTOCORM-LIKE BODY

Nae-Hwey Miao, Yasuko Kaneko and Yasutake Sugawara

Department of Regulation-Biology, Faculty of Science, Saitama University, Saitama, 338-8570, Japan

Abstract

By applying pre-treatment with high concentrations of sucrose and glycerol prior to desiccation and subsequent freezing in liquid nitrogen, successful cryopreservation with high recovery rate was achieved in Oncidium PLB(protocorm-like body). Cellular and subcellular changes after each step in various cryopreservation regimes were examined to elucidate the mechanism of the beneficial effect of the pretreatment which confers resistance to desiccation and freezing.

Key words: Cryopreservation, Desiccation and freezing injury, Electron microscopic observation, Oncidium hamana "elfin", Protocorm-like body (PLB).

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