The Elzone II 5390 Particle Count and Size Analyzer

1. The Elzone II 5390 Particle Count and Size Analyzer Solutions for the Grass Carp Industry

2. Current Situation with Aquatic Vegetation control using Grass Carp • The grass carp, is a large herbivorous fish from lowland rivers eastern Asia. The grass carp, as a biological control of aquatic plants, is considered an attractive long-term method to control the over-growth of submersed aquatic plants. • Because using grass carp to control aquatic vegetation is inexpensive and longer term compared to other control techniques, there is widespread interest in expanding its use throughout the United States. • The diploid grass carp has been used for biological control of aquatic plants since the 1970s. Based on a five-year study, practical guidelines for the use of diploid grass carp to manage aquatic plants in ponds and lakes were developed. However, the potential of the diploid grass carp to naturally reproduce, and eventually over-populate ponds, caused considerable controversy over its use as a biological control agent. This eventually led to the production of non-reproductive, triploid grass carp which most states allow to be used for the control of aquatic plants.

3. Current Situation with Aquatic Vegetation control using Grass Carp • Grass carp, like most fish, are diploid. They possess two sets of chromosomes (one from each parent) and are capable of sexual reproduction. Triploid fish, however, have three sets of chromosomes and are incapable of normal sexual reproduction and the production of viable offspring. The use of triploid grass carp allows benefits of its long-term use for low-cost plant control. • Physical shock techniques induce apparent yields of 100 percent triploids. Because of this high success rate, this technique is being used to produce large numbers of fish. Concern exists, however, that induced polyploidy could result in the production of some reproductive grass carp (i.e., diploid or reproductive triploid fish). Consequently, triploidy in each must be verified prior to stocking. • Ploidy is most often determined by an Electrical Sensing Zone instrument, which electronically measures the volume of a red blood cell after the cell membrane has been chemically removed. Since triploids have larger red blood cells (and nuclei) than diploids, cell size differences are used to confirm triploidy. Larry Sanders, Jan Jeffrey Hoover, and K. Jack Killgore – West Virginia University

4. Elzone II Overview of Basic Theory • Operates using the Electrical Sensing Zone (ESZ) Principle, also known as the Coulter Principle. • First commercially available Elzone system was introduced in 1963. • Fully compliant to the ISO 13319 Standard - Determination of Particle Size Distributions-Electrical Sensing Zone Method, as well as numerous ASTM methods. • Technology has been standardized for use in automated blood cell counters as well as a characterization method for many biological and industrial products. • Thousands of references for the use of various Electrical Sensing Zone models documented. • The highest resolution technology available for particle counting and sizing.

5. Elzone II use for the Grass Carp Industry • The Goal: • Due to the requirement by the U.S. Fish and Wildlife Service to inspect 100% of all Triploid Grass Carp sold, and the fact that there are a limited number of suppliers of Triploid Grass Carp, there is a great need to perform validation tests very quickly (usually a single test should not take more than 6 seconds maximum). • Diploids are approximately 2.8 microns and triploids are slightly larger. Therefore the system used to screen for Triploid and Diploid Carp must have high resolution to discriminate between Diploids and Triploids and be fast enough and able to give real-time results. • The Solution: • Based on the Electrical Sensing Zone method, the Elzone II is capable of making immediate analyses with very high resolution which discriminates Diploids from Triploids. In addition, the operator can screen results immediately using the Elzone II “Sparkle Display.”

6. Elzone II – High Resolution & Real-Time results • The graph shown was done running two latex standards on the Elzone II. One was approximately 2.5microns in diameter (similar to that of a Diploid) and the other was approximately 3 microns in diameter (simulating a Triploid). • Two key results to point out is that the Elzone II has high enough resolution to discriminate between the two standards very well (all the way to the base-line). In addition, you will note that the Sparkle Display will give immediate verification of results. Zoom of the Real-Time Sparkle Display Simulated Diploid pulses as Real-Time as they enter the aperture. Simulated Triploid pulses in Real-Time as they enter the aperture.