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Abstracts: CryoLetters 30 (2), 2009

 

 

Volume 30, No. 2 March/April 2009

ISSN 0143-2044

 

 


MEASUREMENT OF THE MEMBRANE ELASTICITY OF RED  BLOOD CELL WITH OSMOTIC PRESSURE BY OPTICAL TWEEZERS
Jianguang Wu, Yinmei Li, Di Lu, Zhong Liu, Zhengdong Cheng and Liqun He

89-95

 

 


THE EFFECTS OF ELECTRIC FIELD ON ICE NUCLEATION  MAY BE MASKED BY THE INHERENT STOCHASTIC NATURE OF NUCLEATION
P. W. Wilson, K. Osterday and A.D.J. Haymet

96-99

 

 


CRYOPRESERVATION OF ASIAN Dioscorea bulbifera L. AND D. alata L. BY VITRIFICATION: IMPORTANCE OF PLANT GROWTH REGULATORS
Papiya Mukherjee, BB Mandal, KV Bhat and AK Biswas

100-111

 

 


VITRIFICATION OF FARMED BLUE FOX OOCYTES IN  ETHYLENE GLYCOL AND DMSO-BASED SOLUTIONS USING  OPEN-PULLED STRAW (OPS)
Guang-Bin Zhou, Cheng-Bao Ma, Guo-Shi Liu, Shi-En Zhu, Hong-Hai  Zhang, Li-Ling Jia, Lun Suo, Jian-Min Shi, Yong-Bin Wang, Jian-Hui Tian and  Shen-Ming Zeng

112-118

 

 


ANTIFREEZE ACTIVITY OF COLD ACCLIMATED JAPANESE RADISH AND PURIFICATION OF ANTIFREEZE PEPTIDE
Hidehisa Kawahara, Atsuko Fujii, Michihiro Inoue, Satoshi Kitao, Jyoichi Fukuoka and Hitoshi Obata

119-131

 

 


CRYOPRESERVATION OF ZEBRAFISH (Danio rerio)  BLASTOMERES BY CONTROLLED SLOW COOLING
C. Lin, T. Zhang and D. M. Rawson

132-141

 

 


GERMINATION AND EMBRYO RESCUE FROM Passiflora SPECIES SEEDS POST-CRYOPRESERVATION
M. Elena González-Benito, Noemí Aguilar and Teresa Ávila

142-147

 

 


EFFECTS ON CELL VIABILITY OF THREE ZEBRAFISH TESTICULAR CELL OR TISSUE CRYOPRESERVATION  METHODS
C. Bono-Mestre, J. Cardona-Costa and F. García-Ximénez

148-152

 

 

 

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CryoLetters 30 (2), 89-95 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

MEASUREMENT OF THE MEMBRANE ELASTICITY OF RED  BLOOD CELL WITH OSMOTIC PRESSURE BY OPTICAL  TWEEZERS

Jianguang Wu1, 2, Yinmei Li1, 2, Di Lu3, Zhong Liu4, Zhengdong Cheng5 and Liqun He3*

1Department of Physics, University of Science and Technology of China, Hefei 230026, China
2Hefei National Laboratory for Physical Sciences at Microscale, Hefei 230026, China
3Department of Thermal Science and Energy Engineering, University of Science and Technology of China, Hefei 230027, China
4The Red Cross Blood Center of Hefei, Hefei, 230022, China
5Artie MeFerrin Department of Chemical Engineering, Texas A&M University College Station, Texas 77843-3122, USA
*Correspondence author e-mail:
heliqun@ustc.edu.cn

Abstract

Cells have to undergo many changes in osmotic pressure during their long-term preservation, and will have injuries before they return to their normal states. Mechanics of a cell with deformation, either small or large, is usually used to describe the change of the cell quantitatively. However, there are few reports on the deformation of cells subjected to the change of osmotic pressures during preservation. Here, we report our study of the elasticity of the human red blood cells under osmotic pressures using optical tweezers. We find that the deformation characteristics of erythrocytes are strongly dependent on the osmotic pressure. We also find the RBCs will become stiff with increasing osmotic pressure, suggesting a potential reason for membrane injury during preservation.

Keywords: optical tweezers, red blood cell, osmotic pressure, volume change

 

 

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CryoLetters 30 (2), 96-99 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

THE EFFECTS OF ELECTRIC FIELD ON ICE NUCLEATION  MAY BE MASKED BY THE INHERENT STOCHASTIC NATURE OF NUCLEATION

P. W. Wilson*, K. Osterday and A.D.J. Haymet

Scripps Institution of Oceanography, UC San Diego, 9500 Gilman Drive, CA 92093, USA. *Correspondence author e-mail: pwilson@ucsd.edu

Abstract

We use an automatic lag time apparatus to show that an electric field of 5*104 Vm-1 appears to have no effect on the nucleation of supercooled water. Previously reported effects at similar magnitude fields are most likely due to the inherent stochastic nature of liquid to solid nucleation.

Keywords: nucleation, electric field, automatic lag time apparatus, heterogeneous

 

 

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CryoLetters 30 (2), 100-111 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

CRYOPRESERVATION OF ASIAN Dioscorea bulbifera L. AND D. alata L. BY VITRIFICATION: IMPORTANCE OF PLANT GROWTH REGULATORS

Papiya Mukherjee1, 4, BB Mandal1, 5*, KV Bhat2 and AK Biswas3, 6

1Tissue Culture and Cryopreservation Unit, NBPGR, Pusa Campus, New  Delhi-110012, India.
2NRC on DNA Fingerprinting, NBPGR, Pusa Campus, New Delhi-10012, India.
3Department of Botany, University of Kalyani, Kalyani-741235, Nadia, West Bengal, India.
4Agharkar Research Institute, G.G. Agharkar Road, Pune-411004, Maharastra, India.
5A-2, Parijat Apartments, Plot-28, Sector-4, Dwarka, New Delhi-110075, India.
6B 15/93, Kalyani-741235, Nadia, West Bengal, India. 
*Correspondence author e-mail:
mandalbinay@yahoo.co.in

Abstract

The aim of this study was to develop cryopreservation protocols for Asian races of Dioscorea bulbifera and D. alata with high survival and plant regeneration after cryopreservation. Using a vitrification procedure, survival of shoot tips postcryopreservation of up to 89% in D. bulbifera and up to 82% in D. alata were recorded when excised shoot tips were pretreated overnight with 0.3 M sucrose in MS medium, followed by loading with 2 M glycerol plus 0.4 M sucrose for 20 min at 25oC, exposure to PVS2 solution for 90 min at 0oC, immersion in liquid nitrogen for 1 h, rewarming at 40oC for 2 min, unloading in medium with 1.2 M sucrose for 20 min and culturing on growth recovery medium. During growth recovery, 58% shoot regeneration was obtained in D. bulbifera when cryopreserved shoot tips were initially cultured for 40 days on MS medium with 1.5 mg/L BAP, 0.15 mg/L NAA and 0.2 mg/L GA3 followed by culturing on a medium with 0.05 mg/L BAP and 0.15 mg/ L NAA. However, a maximum of 39% shoot regeneration was recorded in D. alata when cryopreserved shoot tips were initially cultured for 40 days on medium M2 (MS containing 1/5 NH4NO3 and 40 g/L sucrose) supplemented with 1.0 mg/L BAP, 1.0 mg/L zeatin, 0.15 mg/L IAA and 0.2 mg/L GA3. Subsequently, the regenerating shoots were cultured for 30 days on medium M2 with 1.0 mg/L BAP, 0.3 mg/L zeatin, 0.02 mg/L NAA and 0.2 mg/L GA3 followed by culturing for another 30 days on medium with 0.5 mg/L BAP, 0.02 mg/L NAA and 0.2 mg/L GA3. Finally, transfer onto medium with 0.05 mg/L BAP and 0.15 mg/L NAA stimulated production of fully grown plantlets. Alteration of post-thaw culture media with plant growth regulators and their application at various stages of growth recovery was crucial for regeneration of shoot tips and formation of plantlets in D. alata.

Keywords: Dioscorea bulbifera, D. alata, cryopreservation, plant growth regulators,
encapsulation-dehydration, vitrification.

 

 

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CryoLetters 30 (2), 112-118 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

VITRIFICATION OF FARMED BLUE FOX OOCYTES IN  ETHYLENE GLYCOL AND DMSO-BASED SOLUTIONS USING OPEN-PULLED STRAW (OPS)

Guang-Bin Zhou1,2,a, Cheng-Bao Ma1,a, Guo-Shi Liu1,*, Shi-En Zhu1,*, Hong-Hai Zhang3, Li-Ling Jia1, Lun Suo, Jian-Min Shi, Yong-Bin Wang, Jian-Hui Tian1 and Shen-Ming Zeng1

1College of Animal Science and Technology, State Key Laboratories for Agrobiotechnology, China Agricultural University, Beijing 100193, P.R. China;
2Dujiangyan campus of Sichuan Agricultural University, Dujiangyan 611830, P.R. China
3Department of Biological Sciences, QuFu Normal University, QuFu 273165, P.R. China
aThese authors contribute equally to the work.
*Corresponding authors email: G.S. Liu (
gshliu@cau.edu.cn);
S.Zhu (
zhushien@ cau.edu.cn)

Abstract

Farmed blue fox was used as a model to develop cryopreservation protocol for nondomestic canine species. We report here the developmental potential of farmed blue fox oocytes after vitrification with a two-step OPS method. Oocytes were collected and pre-cultured for 0, 24, 48, 72 hours respectively before cryopreservation. Vitrification of oocytes was achieved by a 30 sec treatment in 10% ethylene glycol (EG) or 10% EG + 10% dimethyl sulfoxide (DMSO) at 25°C followed by a 25 sec equilibration in EFS30 (30% (v/v) EG +21% (w/v) Ficoll +0.35M sucrose) or EDFS30 (15% (v/v) EG +15% (v/v) DMSO +21% (w/v) Ficoll +0.35M sucrose), before plunging into liquid nitrogen. The survival of oocytes after vitrification was assessed morphologically immediately after warming, and cultured for in vitro maturation. For comparison, control oocytes were cultured for in vitro maturation for 96 hours. The best result was obtained when oocytes were pre-cultured for 72 hours, first exposed to 10% EG + 10% DMSO and vitrified in EDFS30. The survival percentage of oocytes under these conditions was not significantly different (P > 0.05) from that of the control.

Keywords: blue fox (alopex lagopus), oocyte, vitrification; open-pulled straw

 

 

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CryoLetters 30 (2), 119-131 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

ANTIFREEZE ACTIVITY OF COLD ACCLIMATED JAPANESE RADISH AND PURIFICATION OF ANTIFREEZE PEPTIDE

Hidehisa Kawahara1*, Atsuko Fujii1, Michihiro Inoue1, Satoshi Kitao2, Jyoichi  Fukuoka3 and Hitoshi Obata1

1Department of Life Science and Biotechnology, Faculty of Chemistry, Materials and Bioengineering, Kansai University, 3-3-35 Yamate-cho, Suita-shi, Osaka 564-8680, Japan
2Department of Food Science and Nutrition, Faculty of Liberal Arts, Osaka Shoin Women's University, 4-2-26 Hishiyanishi, Higashiosaka-shi, Osaka 577-8550, Japan
3BICC World Co. Ltd., 689-1 Yatabe, Kodera-cho, Himeji-shi, Hyogo 679-2155, Japan.
*Correspondence author
kawahara@ipcku.kansai-u.ac.jp

Abstract

Japanese radish tuber and leaf produced antifreeze proteins (AFPs) having thermal hysteresis activity (TH) and ice recrystallization inhibiting activity (RI). Upon cold acclimation, the apoplastic fluid of the Japanese radish exhibited hexagonal crystal growth, indicating the presence of an antifreeze protein. The induction patterns of protein and the TH activity of apoplastic fraction from both samples were different. The TH activities of apoplastic fraction from tuber and leaves were 0.20±0.03 and 0.18±0.02oC, respectively. Also, the TH and RI activities of apoplastic fraction of leaves were activated by autoclave treatment at pH 10.0. An antifreeze peptide (molecular weight 1,320), was purified using chromatography. Furthermore, the chitinase and β-1, 3-glucanase activities in the apoplastic fraction of its tuber were induced by the cold acclimation. Some proteins in this apoplastic fraction were reacted with the anti-glucanase-like protein (GLP) antiserum and anti-chitinase-like protein (CLP) antiserum produced against isolated winter rye AFPs. This is the first report on the presence and characterization of AFPs from Japanese radish tuber.

Key words: antifreeze protein; Japanese radish; cold acclimation

 

 

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CryoLetters 30 (2), 132-141 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

CRYOPRESERVATION OF ZEBRAFISH (Danio rerio)  BLASTOMERES BY CONTROLLED SLOW COOLING

C. Lin, T. Zhang and D. M. Rawson*

LIRANS Institute of Research in the Applied Natural Sciences, University of Bedfordshire, 250 Butterfield, Great Marlings, Luton, Bedfordshire. LU2 8DL, UK
*Corresponding author email:
david.rawson@beds.ac.uk

Abstract

Cryopreservation is now a common practice in the fields of aquaculture, conservation and biomedicine. One approach for maintaining the genetic diversity of both nuclear genome and mitochondrial DNA is cryopreservation of the blastomere. This study sets out to determine an optimum cryopreservation protocol for blastomeres isolated from 50% epiboly zebrafish embryos. Freezing was performed in 0.25 ml straws in a programmable freezer. The optimum slow cooling protocol is identified as 5°C/min, from 22 to -6ºC, holding for 15 min, 0.3°C/min from -6 to -40°C, 2°C/min from -40 to -80°C, cells were held for 10 min at -80°C before plunging into the liquid nitrogen. Thawing was performed in a water bath at 28°C for 15 s followed by four step-wise removal of the cryoprotectant. Blastomeres had the highest survival level (70.2 ± 3.2%) when a mixture of 1.5 M dimethyl sulfoxide (DMSO) and 0.1 M sucrose were used as cryoprotectants. This was higher than that achieved with either of the sugar alternatives, trehalose (60.6 ± 3.1%) or glucose (43.1 ± 4.9%). In the present study, cryopreservation of 50% epiboly zebrafish blastomeres was studied for the first time using controlled slow cooling method.

Keywords: controlled slow cooling, zebrafish blastomere, sucrose, trypan blue, DMSO.

 

 

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CryoLetters 30 (2), 142-147 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

GERMINATION AND EMBRYO RESCUE FROM Passiflora SPECIES SEEDS POST-CRYOPRESERVATION

M. Elena González-Benito1*, Noemí Aguilar2 and Teresa Ávila2

1Dpto. de Biología Vegetal, Universidad Politécnica de Madrid, 28040 Madrid,  Spain.
2Centro de Investigaciones Fitoecogenéticas de Pairumani, Bolivia.
*Corresponding author
me.gonzalezbenito@upm.es

Abstract

Seeds of Passiflora species have been reported to have intermediate or orthodox storage behavior. The development of cryopreservation protocols for recalcitrant or intermediate seeds can provide a feasible way for long term germplasm conservation. Seed germination of three Passiflora species (P. pinnatistipula, P. tarminiana and P. mollissima) was studied after desiccation and cryopreservation. The three species showed an intermediate response to desiccation: at 3-3.5% water content their germination was reduced to 23-45%, with P. pinnatistipula showing the maximum desiccation tolerance (62% germination after drying to 4.2% water content). The safest seed water contents for cryopreservation: 4.5-4.8% for P. pinnatistipula and P. mollissima, and 9% for P. tarminiana, resulting in 84%, 73%, and 63% germination, respectively. This is the first report of seed cryopreservation for these species.

Keywords: cryopreservation, desiccation, Passiflora, seed, water content

 

 

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CryoLetters 30 (2), 148-152 (2009)
© CryoLetters, c/o University of Bedfordshire, Luton LU2 8DL

EFFECTS ON CELL VIABILITY OF THREE ZEBRAFISH TESTICULAR CELL OR TISSUE CRYOPRESERVATION  METHODS

Bono-Mestre, C1*, Cardona-Costa, J2 and García-Ximénez, F1

1Laboratory of Animal Reproduction and Biotechnology (LARB-UPV), Polytechnic University of Valencia, Camino de Vera 14, 46022 Valencia, Spain.
2Aquaculture and Environmental Research Group (ACUMA), Polytechnic University of Valencia, Camino de Vera 14, 46022 Valencia, Spain.
*Correspondence author email:
bono-mestre@hotmail.es

Abstract

In this work, three cryopreservation procedures were tested in order to obtain efficiently viable testicular cells after cryopreservation.

Testicular cells of Wild type zebrafish males were frozen using an equilibrium protocol and testicular tissue fragments were cryopreserved with equilibrium freezing and vitrification procedures. Results showed that vitrification was significantly more efficient than freezing in terms of final cell survival (cell freezing: 14.4%, tissue freezing: 77.4%-85.5%, tissue vitrification: 94%). It must be noted that, in live cells, the presence of pseudopodia was frequently observed, which indicated their spermatogonial nature.

Based on these results, the authors suggest that vitrification, with the subsequent elimination of connective tissue after warming, offers the best combination to rescue live testicular cells as a genetic conservation procedure in zebrafish.

Keywords: cryopreservation, vitrification, testicular tissue, testicular cell, zebrafish.

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