mesoSPIM_preparing_test_samples

CLARITY test samples

• prepare refractive index matching solution (RIMS) according to your preferences
• use 1 μm beads (e.g. Fluoresbrite YG )
• use agarose (e.g. Sigma-Aldrich 9012-36-6 )
• prepare a 1% agarose solution
• prepare a dilution series of beads in agarose in test tubes, dissolve the beads while the agarose is still hot
• use 10 ml syringes with cut-off tips to pour the agarose. Use the plunger to remove the agarose cylinders after curing
• put the agarose cylinders in RIMS (prepared at an index of nD=1.45) and equilibrate them in an imaging cuvette for at least 24h
• bead samples can be kept for a long time at 4°C

Preparation for first light

• Install MesoSPIM control software and set up config file.
• Set the ETL offsets to roughly the correct position: Run a live acquisition with the scanners turned off. In the ETL tab set the amplitude of the motion to zero with the button. Change the offset value until the focus is mid way between the two objectives. Repeat for the other objective.