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Counting cells in "counting chambers" is a time-consuming process that is prone to error. Scepter accelerates this process and provides precise resultsStage Image

Counting cells in "counting chambers" is a time-consuming process that is prone to error. Scepter accelerates this process and provides precise results

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On the trail of Alzheimer’s

Little angels and little devils

2012/10/15

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The biochemist Kristina Endres has discovered a drug that might benefit dementia patients, although it was originally developed to treat a skin disease. This scientist is conducting vital research at the University of Mainz using Scepter, the first automatic cell counter shaped like a pipette.

  • Kristina Endres of Clinic for Psychiatry and Psychotherapy at Johannes Gutenberg University in Mainz
  • Kristina Endres of Clinic for Psychiatry and Psychotherapy at Johannes Gutenberg University in Mainz
    © PR

    When Kristina Endres explains her daily work she can’t avoid using some cumbersome technical terms. Two words she uses especially often are “alpha-secretase” and “beta-secretase.” These two enzymes compete with each other to cleave a certain protein within the human brain. In a healthy individual their activity is balanced. But sometimes the balance changes in favor of beta-secretase. That person develops Alzheimer's and begins to experience memory loss.

    “Well, there’s a metaphor for what happens,” says Endres. “Basically these enzymes work like little angels and little devils.” Alzheimer's now ranks alongside cancer as one of the world’s most widely researched diseases. In terms of statistics, the reason for the increased incidence is obvious: People keep getting older, and the risk of developing dementia increases with age. But from a scientific point of view, important questions still remain unanswered. What are the root causes of Alzheimer’s disease? And how can it be cured?

    The research lab and the hospital are close together


    Kristina Endres too doesn’t know yet whether she has found the answer. For one thing, the little-angel-and-devil model hasn’t yet been validated in practice. But she intends to change that. “We’re searching for active ingredients that increase the proportion of alpha-secretase without increasing the level of beta-secretase,” she explains. Endres, who is now 37, has been researching the causes of human dementia for 11 years.

    Three years ago she transferred to the Clinic for Psychiatry and Psychotherapy at Johannes Gutenberg University in Mainz to establish and manage the university’s biochemical laboratory. She considers the close proximity of the research lab to the workaday hospital world a great advantage — for instance, it makes it possible to conduct important research projects at close range. “Here we can dovetail basic research and clinical practice,” she says. If everything goes as planned, this close connection will benefit many Alzheimer’s patients in the future. 


    That’s because the meticulous research and many years of experience of Endres and her team have culminated in the discovery of a medication based on a substance that might not only stop the gradual memory loss but perhaps even cure it. Originally this drug, which is similar to a vitamin, wasn’t developed for Alzheimer's at all but instead to treat the skin disease psoriasis. The new discovery benefits from the fact that this drug is already on the market and has therefore already undergone the costly approval process.

    Precise testing of cell cultures


    Tests of the effectiveness and the side effects of the drug are presently under way in Mainz. If the suspected curative potential is confirmed, further tests will be conducted with larger numbers of patients. But Endres won’t make a definitive statement as to whether or when there will be an effective drug against Alzheimer’s. “All the same, there aren’t many research teams pursuing the same approach as we are,” she says. Encouraged by their discovery, the scientists in Mainz are now systematically exploring the pharmaceutical market in search of other drugs that might be useful in treating Alzheimer's. So their typical workday is a lot like searching for the proverbial needle in a haystack.

    “Scepter standardizes the cell-counting process.“

    Kristina Endres
    biochemist
    Johannes Gutenberg University in Mainz, Germany

    Initially the search is conducted “in silico” – with computer programs. Promising active ingredients are then tested “in vitro” in the laboratory. These tests must be designed to be as efficient and comparable as possible, since they are very time-consuming and must be performed repeatedly to ensure valid results. Cell cultures must be handled with particular care. During the early test phases they are not obtained directly from the human brain but as a commercially available, standardized cell culture that has similar properties.  


    After the drugs have been allowed to act on a cell culture for several days, a test is conducted to determine whether and to what extent there has been any increase in alpha-secretase. This information is revealed to the researchers by “messenger substances” admixed with the cells in a liquid. Depending on the amount of alpha-secretase present, these messenger substances fluoresce more or less brightly, and this emitted light can be measured with appropriate instruments. 

    To ensure the validity of these tests, the ratio of messenger substances to cells must be the same from one sample to the next. To standardize such tests, scientists have been using counting chambers for many years. Such a chamber essentially consists of two transparent plates that compress the liquid sample between them into a very fine film. The cells inside this defined space can be counted individually with the help of a microscope and the total number can then be extrapolated for the sample as a whole, which encompasses a few microliters.  


    This process does, however, have some disadvantages. “The measurement process takes more than ten minutes,” says Endres, and she also notes that the result depends to a considerable extent on the employees’ experience and performance level on that particular day. “The measurements are therefore error-prone and often need to be repeated. After all, it makes a huge difference whether I’ve got 45,000 or 60,000 cells per sample,” she explains.

    Thanks to Scepter, scientists no longer have to count cells under a microscope. The convenient handheld device is packed full of technology and supplies fast, precise, and reliable results

    Thanks to Scepter, scientists no longer have to count cells under a microscope. The convenient handheld device is packed full of technology and supplies fast, precise, and reliable results

    © EMD Millipore

    Substances as precious as gold


    She therefore has no regrets about having invested in Scepter – which, according to the manufacturer EMD Millipore, is the first automatic cell counter configured like a pipette. This handy instrument can be operated with a single hand, like a stopwatch. The user's thumb pressure starts an electronic pump that causes a defined volume of the cell suspension to be drawn into a microchannel. A special measuring device determines the number of cells on the basis of the change in electrical resistance.

    The cell size can be determined by extrapolation of the displaced volume. An integrated minicomputer subsequently analyzes the measured values and shows them on the Scepter’s display. This display also makes it possible to compare the results visually and therefore to make swift comparisons. Up to 72 diagrams can be stored directly on the device or transferred to a PC through a USB interface.

    In addition to the convenient operation and analysis, Kristina Endres is particularly impressed by the automation. “With Scepter, the counting process is standardized and takes less than a minute even for beginners," she explains. “My graduate and doctoral students can now make better use of their time.” She adds that this increases the throughput and lowers the error rate. Moreover, experience has shown that a single measuring device is sufficient for the entire biochemical laboratory, where four scientists are usually at work.

    The acquisition has also paid off financially, Endres emphasizes. “Some of the substances we are using are as costly as gold,” she says. “So it shows up on the bottom line very fast if we manage to reduce the number of failed tests.” Other promising substances are available only in limited amounts; they include extracts from rare medicinal plants that staff members bring back from their overseas travels. Such resources are few but potentially important. “Perhaps the solution for combating Alzheimer’s may be growing in Asia or South America,” says Kristina Endres.

    2012/10/15

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    Scepter: Using electrical resistance to count cells

    Scepter is an automatic device for counting cells that are between three and 36 micrometers in diameter. It automates the counting process and accelerates it from over ten minutes to just 30 seconds. The counting is performed by measuring electrical resistance in accordance with the Coulter Principle, which was developed by the American electrical engineer Wallace A. Coulter in 1947. The higher the resistance, the higher the cell concentration of a solution.

    In this process a cell suspension is mixed with an electrically conductive liquid and spread in the gap between two chamber plates. This gap is not much wider than the size of a cell. Ideally the cells, which have high electrical resistance, must traverse this gap one by one – like people in a turnstile. When only the electrically conductive liquid but no cell passes the counter, the resistance is low. But when a cell passes, the resistance increases and this change in resistance can be measured as an electric-current pulse or voltage pulse. That makes it possible to determine (or deduce) the size and number of cells.

     
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