Cell culture robots

Directed evolution relies on the analysis of large numbers of clones to enable the discovery of rare variants with unproved function. In order to analyze these large libraries, methods of screening or selection have been developed, many of which use specialized equipment or automation. These range from the use of multichannel pipettes, all the way up to robotics, depending on the level of investment [59]. Specialized robotic systems are available to perform tasks such as colony picking, cell culture, protein purification, and cell-based assays. 

Figure 3 (A) Robot system for lipofection screening (A) Worktable with racks for microplates, buffer reservoirs, plastic, and glass vials. (B) Four tip liquid handling arm. (C) Gripper for transport of microplates and glass test tubes. (D) High power water bath sonicator. ( ) Nitrogen evaporator. (F) Microplate washer. (G) Absorbance reader. (H) Luminescence reader. (/) Transparent hood. (/) CO2 incubator with pneumatic door (from the rear, front view in B). (B) Self-constructed robotic conveyor for the transport of cell culture plates from the incubator to the worktable.

COS-7 or CHO cells (for initial transfection screening) or cells of therapeutic interest (e.g., dendritic cells and various cancer cells) at a confluence of 50%, grown in 96-well culture plates, were placed into the robot by the robotic conveyor. In a fully automated process, the robot removes the lid from the cell culture microtiter plate, dispenses lipoplexes into the wells (triplicates), replaces the lid and returns the plate to the incubator. After four hours, the cells are automatically retrieved, the cell monolayers are carefully washed using a special drop mode of the integrated plate washer, fresh medium is added, and the cells are incubated for further 42 hours before harvesting. 

Other primary cells, like cardiomyocytes or chondrocytes, can rapidly undergo de-differentiation to fibroblast-like cells when plated on a two-dimensional (flat) surface. In this case, three-dimensional matrices can help maintain the original features. BioLevitator by Hamilton Robotics, 3D Insert by 3dbiotek, and 3D cell culture by Invitrogen are only some of the products available to facilitate the correct growth and morphology of particular primary cell types. 

By definition, the experimental unit is the smallest unit randomly allocated to a distinct level of a treatment factor. Note that if there is no randomization, there is no experimental unit and (in nearly all cases) no experiment. Although it is possible to perform experiments without randomization, it is difficult to do well, and risky unless the experimental system is very well understood (7). Randomization is important for several reasons. Randomization changes the sources of bias into sources of variation in general, a noisy assay is better than a biased assay. Further, randomization allows estimates of variation to represent variation in the population this in turn justifies statistical inference (standard errors, confidence intervals, etc.). A common practice in cell-culture bioassay is to rotate among a small collection of layouts rather than use random allocation. Whereas rotation among a collection of layouts is certainly better than a fixed layout, it is both possible and practical to use carefully structured randomization on a routine basis, particularly when using a robot. 

To provide meaningful toxicity data to discovery chemists during the early phase of lead identification the test systems must be robust and allow data to be delivered in a timely manner so that the information can be incorporated into the decision-makingprocess. At this early time in discovery, compound availability is usually limited, which means that the in vitro systems employed must have low compound requirements. Moreover, these test systems must be amenable to HTS formats, such as 96- or 384-well cell culture, and technically simple for rapid turnaround and successful integration with robotics platforms. 

Many aspects of transporter assays are labor intensive, from culturing cells, to preparing membrane vesicles, to preparing dosing solutions, to sampling transwells. In order to increase throughput and reduce manual labor and potential repetitive hand injuries, several robotic systems have been developed to reduce these laborious tasks, from cell culture maintenance to liquid handling. 

A robot that dispenses culture media, solvents, cell suspensions, antimicrobial agents, or other liquid reagents into 96-well microtiter plates. 

Fig. 1. The five steps to produce arrays of transfected mammalian cells for high content screening microscopy. 1. Preparation of the transfection solutions on an automated liquid handler. 2. Spotting of the transfection solutions with a spotting Robot, for example, ChipWriter Compact on a cell substrate, for example, Lab-Tek tissue culture dishes. 3. Plating of the cells on dishes with dried transfection solutions. 4. Preparation of samples for functional analysis, for example, immunostaining. 5. Analysis of samples by high content screening microscopy.

An extension of the flask culture principle for giving large unit surfece areas are the Nunc A/S Cell Factories (16). These are polystyrene plates of 632 cm which are available in single, double (1264 cm ), 10 tray (6320 cm ), and 40 tray (25280 cm ) units. Each tray level requires 200 ml medium and can be filled, harvested, etc. in one operation per fectoiy unit. For large scale commercial processes the system can be semi-automated with the Manipulator and Robot System. 

Automation and robotics (Terstegge et al., 2007) could minimize the impact of the last issue, but the static nature of the culture would remain. The use of 3D culture systems that more closely resemble the in vivo environment provides increased surface area for cell adhesion and growth, thus leading to higher cellular concentrations but the mass transfer limitations would also increase. 

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