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Cell Biology JoVE (Open Access) Citable · DOI

LDL Cholesterol Uptake Assay Using Live Cell Imaging Analysis with Cell Health Monitoring

DOI: 10.3791/58564-v
What you'll learn
  • ✓ Set up live cell imaging assays to measure LDL cholesterol uptake in real time
  • ✓ Monitor cell morphology and cytotoxicity during compound screening experiments
  • ✓ Analyze and quantify LDL influx rates using image analysis software
Protocol

Biopharma Insights This protocol provides an efficient approach to measuring LDL cholesterol uptake with real time influx rates using a live cell imaging system in various cell types. This technique provides a platform to screen the pharmacological activity of compounds affecting LDL influx while monitoring for cell morphology and hence potential cytotoxicity.

Difficulty
intermediate
Total time
~4–6 hours (assay setup and live imaging acquisition; analysis concurrent or following)
Model organism
Various mammalian cell lines (HEK293, hepatocytes, or other LDL receptor-expressing cells)
Biosafety
BSL-1

Steps

1
Seed cells in 24-well imaging plate

Plate cells at appropriate density in 24-well plates designed for live cell imaging. Prepare cells to reach desired confluency before assay begins.

▶ 00:36
2
Perform live cell imaging analysis

Acquire time-lapse images of live cells using a high-content imaging system to visualize LDL uptake in real time and monitor cell morphology.

▶ 02:54
3
Configure analysis parameters and software settings

Define image analysis parameters including region of interest, LDL signal detection thresholds, and cell morphology metrics in the analysis software.

▶ 03:51
4
Analyze data and calculate influx rates

Process acquired images and extract quantitative LDL uptake kinetics, generating influx rate measurements and assessing cytotoxicity from morphological changes.

▶ 05:41
5
Interpret results across multiple cell types

Compare LDL cholesterol uptake measurements across various cell lines or treatment conditions to validate assay performance and compound activity.

▶ 07:06
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