Clone screening

Process map

Following cell seeding, the transfected cells—typically CHO-S cells—are screened in static culture phase as settled suspension cells; in this way the cells are growing in their normal liquid media environment and are under no additional selection pressure such as having to adapt to growth in semi-solid media.

For clone screening, image quality is crucial. High-quality images at each stage of the process are necessary for the operator to make confident decisions, streamline the workflow and optimise laboratory productivity.

A clearer view

Image of well edge

Conventional systems for automated cell imaging have been limited by variable image quality and insufficient coverage of the well edges where cells tend to stick and grow. To overcome this, the Cell Metric CLD™ has the following attributes:

  • Advanced optics generate clear, sharp images of adherent or settled suspension cells, showing more detail than previously possible. Easily distinguish between single cells, pairs of cells, and debris. It is possible to see a single cell in the well within minutes of dispensing the cells.
  • Consistent illumination across the whole well, even at the well edge, which is normally in shadow on other imaging systems. This means that all cells are detected by the Cell Metric CLD™. When the software detects a single cell in a well, the operator can be confident of a monoclonal origin.
  • Automatic focusing corrects for any plastic distortion across the plate or individual well and means that every part of every well is perfectly in focus. All cells are clearly seen and documented.

An important factor to also consider in producing the best possible image quality is the type and quality of microplate used. The combination of a high-quality plate and the high-quality images from the Cell Metric CLD™ can give the operator the confidence to make an early clone selection.

Earlier evidence of monoclonality

Ensuring cell lines are monoclonal (derived from a single cell) is a key requirement of cell line development. With traditional approaches cells were checked several days after seeding when a small colony had already formed. By this time it was easier to see the cells but it was only possible to decide whether each well contained a single colony, not a single cell. The top candidate clones were then subcloned again to further reduce the likelihood that more than one cell was deposited, although this did not provide definitive evidence of monoclonality.

In contrast, photo-documentation by the Cell Metric CLD™ is now used to demonstrate monoclonality on the day of seeding, before cells have started to proliferate. This means that subcloning in order to increase the probability of monoclonality is no longer necessary. The instrument has sufficient resolution and sensitivity to clearly identify a single cell in each well within hours of seeding. A permanent digital record is produced as evidence of monoclonality.

Existing Cell Metric CLD™ users have already verified that they have saved many weeks of upstream development time by not having to subclone.

HeLa cells imaged on the Cell MetricLabel-free imaging

The use of fluorescent tags can potentially alter cell growth, proliferation and gene expression, and in some cases can render the final product unacceptable for biotherapeutic use. To overcome this, the Cell Metric CLD™ uses high-resolution brightfield cell imaging, meaning there is no requirement for fluorescent tags or labels for cell screening. This brings you closer to the in vivo environment, which is important where the ultimate cell line will be used in the production of biotherapeutic medicines.

Accurate workflow management

A typical cell line development project may involve tens to hundreds of plates seeded with cells. The exact number of plates depends on the technology used, the number of wells per plate (384 or 96 wells) and the occupancy rate. Previously these plates would have been laboriously viewed by eye, using a microscope to check each well and determine how many cells were present. A manual record of the data would then have been created.

The Cell Metric CLD™ now completely automates this task and stores a clear image of the whole of each well to provide unequivocal evidence that a well contained a single cell. After scanning, wells containing single cells are highlighted by the software, for easy identification and tracking. This process drastically reduces operator input and removes subjectivity as well as user error. Clone documentation including the image history of every single well is retained so you can track from colony back to single cell. In addition, the ability of the Cell Metric CLD™ with its automated microplate loader to operate unattended frees up laboratory personnel for more valuable tasks.

Colony growth from a single cell

The next step

The most promising clones are next assessed for cell growth and stability.

To find out more, go to: