PTM Based Tumor Microenvironment Monitoring: A Method for Tracking Changes in ECM Rigidity Using Tumor Spheroids

Published: April 30, 2023

Abstract

Source: Jones, D. P., et al. Longitudinal Measurement of Extracellular Matrix Rigidity in 3D Tumor Models Using Particle-tracking Microrheology. J. Vis. Exp. (2014).

This protocol describes a technique for monitoring extracellular matrix rigidity changes by using particle-tracking microrheology (PTM). This is done by culturing 3D pancreatic tumor spheroids into ECM with embedded fluorescent probes for video particle-tracking and analysis.

Protocol

1. Preparing 3D Tumor Spheroids Embedded in ECM Prepare a work station with needed materials inside a laminar flow hood. Prepare a diluted mixture of carboxylate-modified 1 μm diameter fluorescent tracer probes by adding 2 parts stock probes (2% solids) to 25 parts sterile water. Remove a bottle of 3.1 mg/ml bovine collagen from the refrigerator and place it on ice. Aliquot 125 μl of collagen into an empty 2 ml vial. Add 50 μl of diluted …

Açıklamalar

The authors have nothing to disclose.

Materials

Bovine type 1 collagen BD Biosciences, San Jose, CA 354231
PANC-1 American Type Cell Culture, Manassas, VA CRL1469 or other appropriate cell type
Fluorescent Microspheres Life Technologies, Carlsbad, CA 906906
Agarose Fisher Bioreagents, Waltham, MA C12H18O9
NaOH Fisher Bioreagents, Waltham, MA NC0480985
96-well Imaging plates Corning Inc., Corning, NY 3904
DMEM Hyclone, Waltham, MA SH30243.01 or appropriate cell culture media
Zeiss AxioObsever Microscope Zeiss, Oberkochen, Germany includes high-speed camera and imaging software

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Bu Makaleden Alıntı Yapın
PTM Based Tumor Microenvironment Monitoring: A Method for Tracking Changes in ECM Rigidity Using Tumor Spheroids. J. Vis. Exp. (Pending Publication), e20323, doi: (2023).

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