Optical Imaging

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We use calcium dyes to measure the activity of neurons during seizures.

Staining techniques we use are for Juvenile and Adult Acute Cortical Slices. Details are given in Chapter 43 by Ross, Nakamura, Watanabe, Larkum and Lasser-Ross from: Imaging in Neuroscience and Development: A laboratory manual. Eds. Yuste and Konnerth, Cold Spring Harbor Press.

They recommend staining the slices with a bolus of dye dissolved in DMSO and Pluronic acid. The DMSO is to help dissolve the membranes a little to get the dye into it, while the Pluronic Acid is supposed to prevent the dye from being taken up into the organelles, a process called compartmentalization.

Here is our recipe: Oregon Green BAPTA 2: MW 1751.45 g/mol at 50ug Pluronic acid: 2 uL DMSO: 26 uL or 24 mg

Final Volume = 50e-6g/(1751.45g/mol*1e-3mol/L)=28uL

in 2.5cc of oxygenated ACSF add 7uL of dye and let incubate for 20-30 min at physiological temperatures.

Maximum Settings:

25 Hz 256 BNC/CCD ratio 8192 frames with 1 BNC recording

25 Hz 128 BNC/CCD ratio 16384 frames with 1 BNC recording


Getting Started With Imaging

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