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Related Experiment Video

Updated: Nov 11, 2025

A Versatile Method of Patterning Proteins and Cells

A Versatile Method of Patterning Proteins and Cells

Published on: February 26, 2017

Patterning Neuroepithelial Cell Sheet via a Sustained Chemical Gradient Generated by Localized Passive Diffusion

Ningwei Li¹, Feiyu Yang¹, Subiksha Parthasarathy²

¹Department of Mechanical and Industrial Engineering, University of Massachusetts Amherst, Amherst, Massachusetts 01003, United States.

ACS Biomaterials Science & Engineering

|March 22, 2021

Summary

Localized passive diffusion (LPaD) devices create stable chemical gradients for patterning human pluripotent stem cells (hPSCs) in vitro. This method simplifies gradient generation for studying early human development without complex equipment.

Keywords:

chemical gradient human pluripotent stem cells microfluidics neuroepithelium patterning

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Published on: February 16, 2012

Area of Science:

Stem cell biology
Developmental biology
Bioengineering

Background:

Human pluripotent stem cells (hPSCs) offer novel in vitro models for early human development.
Current methods for patterning neuroectoderm tissues in vitro using morphogen gradients face limitations like complex equipment and downstream analysis challenges.

Purpose of the Study:

To develop a simplified method for generating stable chemical gradients in vitro.
To assess the efficacy of localized passive diffusion (LPaD) for patterning hPSC-derived tissues.

Main Methods:

A novel device utilizing localized passive diffusion (LPaD) was designed, featuring a gelatin-filled slit connected to a morphogen reservoir.
Chemical gradient stability was assessed using DAPI fluorescent intensity over 168 hours.
The device's ability to induce cellular responses was tested with Madin-Darby canine kidney (MDCK) cells and cytochalasin D, and for patterning hPSC-derived forebrain neuroepithelial cells with sonic hedgehog (Shh).

Main Results:

LPaD devices successfully generated stable chemical gradients for up to 168 hours in an open environment.
The devices induced specific cellular responses in MDCK cells to a cytochalasin D gradient.
Efficient patterning of the dorsal-ventral axis in hPSC-derived neuroepithelial cells was achieved using an Shh gradient generated by LPaD devices.

Conclusions:

LPaD devices provide a simple, stable, and adaptable method for generating chemical gradients in vitro, overcoming limitations of microfluidic systems.
These devices are effective tools for controlling the local chemical microenvironment, enabling the engineering of organotypic structures and facilitating the study of developmental processes.