Protocol for in vitro imaging of molecular assemblies

Mounting method for in vitro imaging of molecular assembly using CherryTemp

This protocol shows how to create a chamber on a glass coverslip to image in vitro biochemical reactions.

Placing a CherryTemp chip on top of this chamber allows for thermalization of the sample, to observe temperature dependent molecular dynamics in the seconds time scale.

Two types of spacers can be used:

-LSR elastomere: for volumes up to 10ul
-Silicone spacer: to create a sealed chamber for larger volumes up to 50µL


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Step 1: Remove the protection from the dedicated spacer (in vitro applications, red spacer) and paste it on top a 24x24mm glass coverslip.

fig 1 in vitro

Step 2: Remove the second protection from the silicone spacer.

fig 2 in vitro

Step 3: Pipette your sample on the coverslip.

Fig 3 in vitro

Step 4: Top the glass coverslip with the CherryTemp chip, by sticking it to the spacer. Place the chip with the correct orientation (thermalization chamber facing down, in contact with the spacer), with the thermalization chamber centered on the sample.

fig 4 in vitro

Step 5: Place the mounted system on either an upright or an inverted configuration, depending on your microscope setup.

fig 5 in vitro

Step 6: When finished, remove the CherryTemp chip from the coverslip

– The CherryTemp chip can be cleaned by wiping the chamber part with ethanol (don’t soak the chip on  ethanol)
– The silicone spacer can be cleaned with ethanol

Protocol adapted from Rosen Lab, University of UT Southwestern Medical Center in Dallas.

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