Immunofluorescence Staining of F-actin



March 7, 2019

EP151 v1.0

 

Goals:

Key Steps:

Other Required Materials:

Visualize F-actin via fluorescent imaging in Emulate Organ-Chips

  • Immunofluorescence staining in chip
  • Alexa Fluor™ 488 Phalloidin (InvitrogenTM A12379)
  • Dimethyl sulfoxide (DMSO)
  • PBS
  • 4% paraformaldehyde (PFA)
  • 10% Saponin
  • PBS
  • BSA
  • Fluorescence microscope

Method

Sample type

Fixed Organ-Chip

See Protocol EP137 Fixation and Immunofluorescence (IF) Staining.

Recommended fixative and incubation time

4% PFA, 15 minutes at room temperature

Recommended permeabilization and incubation time

1% saponin in PBS, 30 minutes at room temperature

Recommended blocking buffer and incubation time

1% BSA,10% goat serum in PBS, overnight at 4°C

Recommended primary antibody dilution and incubation time

Alexa Fluor™ 488 Phalloidin (InvitrogenTM A12379)

Dilute 1:1000 in PBS (stock prepared in DMSO), 30 minutes incubation at room temperature

Representative image

Image of F-actin (green) in liver sinusoidal endothelial cells in the Liver-Chip (bottom channel).

More information on vendor site

https://www.thermofisher.com/order/catalog/product/A12379

 


 

© Emulate, Inc., 2019. All rights reserved.

Emulate® and the Emulate logo are registered trademarks for Emulate, Inc.

InvitrogenTM and Alexa Fluor™ are trademarks of Thermo Fisher Scientific or its subsidiaries.

The technology disclosed in this document may be covered by one or more patents or patent applications owned by or licensed to Emulate, Inc. No license is granted herein. You are solely responsible for determining whether you have all intellectual property rights that are necessary for your intended use of Emulate products or protocols, and whether you are required to obtain any additional intellectual property rights from a third party. Further information is available by contacting Emulate.