Cell counting | EPFL Graph Search

Cell counting is any of various methods for the counting or similar quantification of cells in the life sciences, including medical diagnosis and treatment. It is an important subset of cytometry, with applications in research and clinical practice. For example, the complete blood count can help a physician to determine why a patient feels unwell and what to do to help. Cell counts within liquid media (such as blood, plasma, lymph, or laboratory rinsate) are usually expressed as a number of cells per unit of volume, thus expressing a concentration (for example, 5,000 cells per milliliter). Uses Numerous procedures in biology and medicine require the counting of cells. By the counting of cells in a known small volume, the concentration can be mediated. Examples of the need for cell counting include:

In medicine, the concentration of various blood cells, such as red blood cells and white blood cells, can give crucial information regarding the health situation of a person (

New disposable tubes for rapid and precise biomass assessment for suspension cultures of mammalian cells

David Hacker, Nicolas Jaccard, Martin Jordan, Mathieu Benjamin Stettler, Florian Maria Wurm

We present a new approach for biomass assessment in cell culture using a disposable microcentrifuge tube. The specially designed tube is fitted with an upper chamber for sample loading and a lower 5 microL capillary for cell collection during centrifugation. The resulting packed cell volume (PCV) can be quantitatively expressed as the percentage of the total volume of the sample. The present study focused on the validation of the method with mammalian cell lines that are widely used in bioprocessing. Using several examples, the PCV method was shown to be more precise, rapid, and reproducible than manual cell counting.

2006

An impedance-based flow microcytometer for single cell morphology discrimination

Ludovica Colella, Philippe Renaud, Marjan Shaker

Cell shape is a fundamental biological feature, providing specific information about physiological or pathological cellular conditions. Most of the state-of-the-art microfluidic cytometers, however, only allow simple cell analysis, including viability studies, cell counting and sorting. In this work, we present a non-invasive, label-free device capable of single cell morphology discrimination in continuous flow. The device is based on the principle of liquid electrodes, fabricated in a cross configuration around a sensing zone. This arrangement allows measurement of cell impedance along orthogonal orientations and extraction of an index describing cell shape anisotropy. By adding prior to the sensing volume a series of lateral liquid electrodes, the particle stream was focused toward the channel midline and each cell was oriented in a specific direction before shape sensing. We demonstrate the proof of concept by performing spherical and elongated particle discrimination. As an application, we show that the shape changes experienced during cell division can be monitored and characterized. In particular, budding yeasts at different stages of the mitotic cycle were identified by extracting their anisotropy index.

Royal Soc Chemistry2014

Bioreactor processes based on disposable materials for the production of recombinant proteins from mammalian cells

Mathieu Benjamin Stettler

The objective of industrial cell culture technology is to produce high value-added therapeutic proteins that are well suited for the treatment of infectious diseases, cancer, and autoimmune diseases. Nowadays, an important focus is the reduction of the time and costs from the discovery of the candidate molecule to the full-scale production. Relying on enabling technologies is critical for meeting this objective. Simultaneously, novel technologies are expected to increase the clinical and commercial success rates. In recent years, new concepts based on innovative disposable materials emerged and appeared to be promising alternatives to standard bioprocessing equipment. This thesis work was focused on the use of such disposable compounds in mammalian cell culture applications, particularly for the containment of the cells, and was aimed at demonstrating their benefit in terms of cost-effectiveness, ease-of-use and flexibility. First, a novel disposable packed cell volume tube was shown to be ideal for the quick and reliable assessment of biomasses. A characterization and validation of the measurement system demonstrated that it could replace time-consuming and less accurate cell counting methods. Then, to determine the appropriate growth conditions for cells in culture, small-scale single-use tubes were orbitally agitated with conventional lab shakers. This approach was found to be well-suited for multi-parameter optimization strategies. A systematic characterization of the liquid mass transfer in shake tubes proved that sufficient oxygen was available even at cell densities beyond 1 ×107 cells mL-1. Non-invasive optical methods were used to assess the dissolved oxygen variations in various orbital shake vessels, from the milliliter to the multi-liter scale. This simple and yet powerful cultivation principle, orbital shaking, was scaled-up to pilot and production scales. To match the requirements in terms of oxygen transfer at larger scales, a given airflow was provided to replace the gas in the headspace. When required, the airflow was enriched with oxygen. At scales above 20 L, sterile disposable cell cultivation bags were used to contain the cells. Combining orbital shake technology and disposable cell culture bags with a cylindrical shape was promising, both in terms of efficiency and ease-of-use. Prototype shake bioreactors of 200 L and 1'500 L were designed, constructed and operated with maximal cell densities up to 5-7 ×106 cells mL-1 in batch cultivations of CHO cells. The benefits in terms of time and cost savings were even more obvious when novel shake bioreactors were compared with standard stirred-tank bioreactors. This was shown in a realistic optimization, scale-up and production sequence. Most importantly, this work established that orbital shake technology, unlike other disposable cell cultivation systems, can be used for a wide range of operating scales, from only a few milliliters for optimization purposes up to production scale.

EPFL2007

Bioreactor processes based on disposable materials for the production of recombinant proteins from mammalian cells

Mathieu Benjamin Stettler

EPFL2007