Oct 23, 2024
Low-Cost Microfluidics For Light Microscopy Experiments
This protocol is a draft, published without a DOI.
- 1East Carolina University
Protocol Citation: Cameron Schmidt 2024. Low-Cost Microfluidics For Light Microscopy Experiments. protocols.io https://protocols.io/view/low-cost-microfluidics-for-light-microscopy-experi-dp8t5rwn
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 22, 2024
Last Modified: October 23, 2024
Protocol Integer ID: 110579
Funders Acknowledgement:
Cameron A. Schmidt
Grant ID: Eunice Kennedy Shriver National Institute of Child Health and Human Development (R01HD110170)
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Abstract
Microfluidics devices are powerful tools for studying dynamic processes in
live cells, especially when used in conjunction with light microscopy. There
are many applications of microfluidics devices including recording dynamic
cellular responses to small molecules or other chemical conditions in perfused
media, monitoring cell migration in constrained spaces, or collecting media
perfusate for the study of secreted compounds in response to experimental
inputs/manipulations. Here we describe a configurable low-cost (channel-based)
microfluidics platform for live-cell microscopy, intended to be useful for
experiments that require more precision/flexibility than simple rubber spacers,
but less precision than molded elastomer-based platforms. The materials are
widely commercially available, low-cost, and device assembly takes only minutes.
Prepare the plastic slides from PET plastic sheets
Prepare the plastic slides from PET plastic sheets
Measure the desired slide size on the plastic sheets and mark with a marker.
Score the shapes on both sides of the plastic with a hobby knife using a ruler as a guide.
Use the edge of a table or workbench to snap the shapes along the scored lines. Note: the plastic sheets are covered with a thin liner. Do not remove these until absolutely necessary to avoid getting dust or debris on the plastic.
Once the spacer has been created (see section below), lay the top slide on the spacer to observe the alignment and mark the inflow and outflow points with a marker.
Tape the slide onto a styrofoam block or other soft surface. Wet the points on the plastic with a drop of water. Using a rotary tool and 1mm drill bit, drill holes in the plastic.
Connect the tubing adapters
Connect the tubing adapters
Peel the backing off of the tubing adapter (Grace Biolabs Cat #460003) and attach above the drilled hole in the top slide. Insert a blunt tipped 21.5 gauge needle through the side with the adhesive (so that the needle end comes out of the top).
Attach a (desired) length of of PE-50 tubing (Intramedic Cat. #14-170-12B) onto the blunt tip of the needle.
Draw the needle through to pull the tubing into the tubing adapter. Apply pressure to the tubing adapter when the tubing is approximately 85% of the way through to cause the needle to release, leaving the tube in the adapter.
Repeat for the desired number of inflow/outflow tubing connections.
Prepare the spacer
Prepare the spacer
Using the Cricut Design Space software, create the desired channel layout.
Using scissors or a paper trimmer, cut the double sided adhesive sheet into an appropriate length and width for the Cricut machine. Load the adhesive onto the machine with the top and bottom backings still in place.
Initiate the design cutting process from the Cricut Design Space software.
Loaded Cricut machine.
Using scissors, cut the spacers out of the larger piece of double sided adhesive sheet.
A spacer cut out of the larger adhesive sheet.
Using a seeding tool or any fine pointed tool, remove the top backing and adhesive from the internal regions of the channels.
Spacer with internal portion of the channel space removed.
Spacer with internal portion of the channel space removed.
Peel off the spacer with the top backing still in place and apply to the bottom plastic slide or cover slip.
Peel the top backing off of the adhesive spacer.
Assemble the device
Assemble the device
Align the inflow/outflow adapters with the desired positions on the spacer and carefully place the top slide on the adhesive. This results in a fully assemble device.
Note: Alternatively, the tubing can be threaded into the tubing adapters without pre-attaching them to the top slide. Then, the top and bottom slides can be compressed between two pieces of wood or other material and clamped to form a tighter seal between the slides. The tubing adapters can then be attached. This approach generally results in more reliable seals.
Pressure test
Pressure test
Perform a pressure test by attaching a syringe with blunt tipped needle to the inflow tubing and placing the outflow tubing in a small beaker to collect effluent. Any suitable contrast dye can be used for visualization of the seal integrity. Using a perfusion pump, fill the device and observe for leaks. Note that perfusing by hand may cause leakage due to the high flow rates. Though this may depend on the materials used in the assembly as well as other factors such as temperature.
