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Cell Counting and Image Cytometry

Growth Inhibition

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Topic
Growth Inhibition
Embryoid Bodies and Patient-Derived Organoids
3D Confrontation Assays
Growth Inhibition
Measure Apoptosis
Measure Migration onto extracellular matrix
Quantify Invasion into Matrigel
Tumor Spheroid Viability
Tumorsphere Formation & Clonogenic Survival

Image and measure the size of 3D tumor spheroids under multiple drug and growth conditions over time

  1. Directly image tumor spheroids in various microwell formats
  2. Non-invasive brightfield imaging allows the user to image the same plate over multiple days
  3. Measure spheroid volume and diameter concurrently with spheroid imaging

Introduction

The Celigo™ image cytometer has been developed to fully automate live cell imaging analysis of tumorspheres. This automated morphometric analysis tool significantly reduces the time and effort needed to quantify key aspects of 3D spheres including size, growth, growth tracking over time and response to chemotherapeutics.

growth inhibition celigo image cytometer

Drug-Induced tumor spheroid growth inhibition

growth inhibition experimental setup

Experimental setup


4-day old spheroids were treated with compounds for 72 hours. Drug-induced concentration-dependent growth inhibition was obtained.

  • Day 0-4: form spheroids
  • Day 4-7: drug treatment
  • Day 4-14: measure spheroid size
     
growth inhibition drug concentration

Drug concentration
Green calcein AM stained target cells

growth inhibition green-calcein

U 87-MG (giloblastoma) + 17-AGG
Green calcein AM stained target cells

Live cell analysis of spheroid growth inhibition under drug treatment and hypoxia

Experimental protocol:

  1. Seed 2500 NCI-H460 cells/well in a ULA 96-well plates
  2. On day 4 treat with drug X or vehicle control
  3. Place one plate in hypoxic conditions and one plate at a normoxic condition
  4. On day 4 post-treatment image entire 96-well plate in ~5 minutes

At high drug dose, NCI-H460 3D tumor spheroids showed growth inhibition under hypoxic conditions

 

growth inhibition spheroid at normoxia

NCI-H460 Spheroid at Normoxia, treated with Drug X at 10 µM

growth inhibition normoxia vs hypoxia conditions

NCI-H460 cells treated with Drug X and grown in Normoxia vs Hypoxia conditions

spheroid at hypoxia

NCI-H460 Spheroid at Hypoxia, treated with Drug X at 10 µM

Captured brightfield images of NCI-H460 spheroids treated with a 10 µM drug and grown at different oxygen conditions show that spheroid size is affected by growth conditions.

By imaging the same plate over multiple days, drug-treated spheroids at hypoxic conditions had smaller spheroid diameters compared to the spheroids grown at normal conditions.

For research use only. Not for use in diagnostic procedures.

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