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Portable Cryo-plunger for Cryogenic Microscopy Sampling

IB-3145

APPLICATIONS OF TECHNOLOGY:

ADVANTAGES:

ABSTRACT:

A Berkeley Lab team led by Luis R. Comolli has developed the Portable Cryo-Plunger, a lightweight, portable flash-freezing apparatus that can safely preserve biological samples in the field for subsequent analysis using cryogenic transmission electron microscopy (cryo-TEM), an approach to study near-intact bacteria, viruses, and protein structures if they are flash frozen in vitreous (non-crystalline) ice. The Portable Cryo-Plunger has successfully prepared cryo-TEM samples in real time, enabling the study of microbial communities that have never been cultured in a laboratory setting.

As demonstrated in one configuration of the device, weighing 4 kg and standing 71-cm tall, the Portable Cryo-Plunger can be transported on foot to a sampling site (see image below). Mounted on a tripod, it can be set up in difficult terrain, such as the interior of a mine or the shores of a salt lake. The Portable Cryo-Plunger is designed to fit in its base, a medium density foam cylinder Dewar containing a liquid nitrogen bath, surrounding a liquid ethane or propane cup where the flash freezing (~-188 °C) takes place. A droplet of sample is applied to a standard cryo-TEM grid, and the grid is placed at the plunger tip. At the top of the plunger is a spring-loaded handle that is used to drive the sample tip quickly into the flash-freezing cup.

The use of Cryo-TEM has been limited because most specimens must be flash frozen in a lab, and by the time they reach the lab from remote field locations, they can be seriously degraded. Freezing eliminates damage caused by traditional fixation and dehydration of specimens, affording more realistic and detailed views of bacteria, including images of delicate extracellular structures whose surfaces are exposed to the environment.

Since the Cryo-Plunger costs less than a tenth of the price of commercially available cryo-TEM flash freezing devices, it can be used in laboratories that do not have the resources to acquire automated equipment. The device also can foster collaborations between cryo-TEM labs and field researchers who previously lacked access to cryo-TEM microscopy.

     
  cryoplunger    
 

The Cryo-Plunger is designed for ease of operation in the field. This version, set up on a tripod, weighs only 4 kg.

 

DEVELOPMENT STATUS: Fully functional field-tested prototype. 

STATUS: Patent pending. Available for licensing or collaborative research.

FOR MORE INFORMATION:

Comolli, L.R., Duarte, R., Baum, D., Luef, B., Downing, K.H., Larson, D.M., Csencsits, R., Banfield, J.F. “A portable cryo-plunger for on-site intact cryogenic microscopy sample preparation in natural environment,” Microscopy Research and Technique, December 30, 2011.

Russell, Sabin. “Taking the Cryo Plunge: Flash-freezing in the Field,” Berkeley Lab TechStream, May 16, 2012.

REFERENCE NUMBER: IB-3145

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Last updated: 05/18/2012