36
EVOS
™
M5000 Imaging System User Guide
Measure confluence
Confluence tool
Confluence is a measure of how tightly cells are packed in a culture. When all
available growth area is utilized in a culture vessel and the cells make close contact
with one another, the culture is at 100% confluence.
Guidelines for
confluence
measurements
•
We recommend that you visualize your cells using transmitted lightand a
phase objective with
4X
to
10X
magnification. Set the phase ring to
Oly 4×
(for
Olympus
™
4X phase objective) or
4×/10×
(for EVOS
™
phase objectives) using
the phase annuli selector (page 7).
•
Increasing the number of targets and background areas improves accuracy.
You can select up to 5 target areas and 5 background areas.
•
The Confluence tool uses a texture and intensity based algorithm. The
sensitivity slider adjusts the algorithm sensitivity to pixel intensity (higher
intensity = more pixels included). Decreasing the sensitivity reduces the
confluence value.
•
Different cell types have different confluence “patterns”, and variability in
morphology and contrast can influence the absolute confluence measurement
between different cell types. However, with a given cell type, the reproducibility
from measurement to measurement can be optimized. Reproducibility in
confluence measurements is more important than absolute percentages.
Measure
confluence
1.
In the transmitted light channel, adjust lighting and focus on the sample, then
click
Capture
to acquire an image of your culture.
2.
Hover the pointer over the Viewing area to reveal
Display Settings and
Analysis Tools
, then click the
Analysis
button.
3.
In the tabs area, select
Confluence
to display the Confluence tool.
